Relationship of airway narrowing, compliance, and cartilage in isolated bronchial segments.

Structural components of the airway wall may act to load airway smooth muscle and restrict airway narrowing. In this study, the effect of load on airway narrowing was investigated in pig isolated bronchial segments. In some bronchi, pieces of cartilage were removed by careful dissection. Airway narrowing was produced by maximum electrical field stimulation. An endoscope was used to record lumen narrowing. The compliance of the bronchial segments was determined from the cross-sectional area of the lumen and the transmural pressure. Airway narrowing and the velocity of airway narrowing were increased in cartilage-removed airways compared with intact control bronchi. Morphometric assessment of smooth muscle length showed greater muscle shortening to acetylcholine in cartilage-removed airways than in controls. Airway narrowing was positively correlated with airway compliance. Compliance and area of cartilage were negatively correlated. These results show that airway narrowing is increased in compliant airways and that cartilage significantly loads airway smooth muscle in whole bronchi.     

Biophysical basis for airway hyperresponsiveness

Airway hyperresponsiveness is the excessive narrowing of the airway lumen caused by stimuli that would cause little or no narrowing in the normal individual. It is one of the cardinal features of asthma, but its mechanisms remain unexplained. In asthma, the key end-effector of acute airway narrowing is contraction of the airway smooth muscle cell that is driven by myosin motors exerting their mechanical effects within an integrated cytoskeletal scaffolding. In just the past few years, however, our understanding of the rules that govern muscle biophysics has dramatically changed, as has their classical relationship to airway mechanics. It has become well established, for example, that muscle length is equilibrated dynamically rather than statically, and that in a dynamic setting nonclassical features of muscle biophysics come to the forefront, including unanticipated interactions between the muscle and its time-varying load, as well as the ability of the muscle cell to adapt (remodel) its internal microstructure rapidly in response to its ever-changing mechanical environment. Here, we consider some of these emerging concepts and, in particular, focus on structural remodeling of the airway smooth muscle cell as it relates to excessive airway narrowing in asthma.     

Decreased airway narrowing and smooth muscle contraction in hyperresponsive pigs.

Increased smooth muscle contractility or reduced smooth muscle mechanical loads could account for the excessive airway narrowing and hyperresponsiveness seen in asthma. These mechanisms were investigated by using an allergen-induced porcine model of airway hyperresponsiveness. Airway narrowing to electric field stimulation was measured in isolated bronchial segments, over a range of transmural pressures (0-20 cmH(2)O). Contractile responses to ACh were measured in bronchial segments and in isolated tracheal smooth muscle strips isolated from control and test (ovalbumin sensitized and challenged) pigs. Test airways narrowed less than controls (P < 0.0001). Test pigs showed reduced contractility to ACh, both in isolated bronchi (P < 0.01) and smooth muscle strips (P < 0.01). Thus isolated airways from pigs exhibiting airway hyperresponsiveness in vivo are hyporesponsive in vitro. The decreased narrowing in bronchi from hyperresponsive pigs may be related to decreased smooth muscle contractility. These data suggest that mechanisms external to the airway wall may be important to the hyperresponsive nature of sensitized lungs.     

Structural basis for exaggerated airway narrowing

Airway hyperresponsiveness, particularly the ability of airways to narrow excessively in response to stimuli that normally cause little airway narrowing in nonasthmatic subjects, is a characteristic feature of asthma and the basis of its symptoms. Although airway hyperresponsiveness may be partly the result of alterations in the contractile phenotype of the airway smooth muscle, there is evidence that it may also be caused by structural changes in the airway wall, collectively termed airway remodeling. Airway remodeling is defined as changes in composition, quantity, and (or) organization of cellular and molecular constituents of the airway wall. Airway wall remodeling that occurs in asthma can result in functional alterations because of quantitative changes in airway wall compartments, and (or) because of changes in the biochemical composition or material properties of the various constituents of the airway wall. This brief review summarizes the quantitative changes in the dimensions and organization of the airway wall compartments that have been described and explains how structural alterations may lead to the exaggerated airway narrowing.     

Self-organized patterns of airway narrowing

The behavior of respiratory diseases such as asthma and COPD may involve complicated interactions among multiple factors. Theoretical and experimental data suggest that interdependence among the airways of the bronchial tree leads to the emergence of self-organized patterns of airway narrowing, ventilation defects, and other phenomena when a tipping point is passed. Additionally, evidence from several studies shows that the behavior of an isolated airway is different from an identical airway embedded in the bronchial tree so that experimental results of isolated elements such as airways, airway smooth muscle, or inflammatory pathways may not explain the whole organ behavior. However, there may be factors in the isolated elements that can dramatically change the complex system's behavior. More effective strategies for prevention or recovery from a disease, such as asthma, will depend on our progress in identifying and understanding the essential parts of the self-organized behavior that is involved.     

Blood flow distribution within the airway wall.

Altered perfusion of the bronchial mucosal plexus relative to the adventitial plexus may contribute to geometric changes in the airway wall and lumen. We studied bronchial perfusion distribution in sheep by using fluorescent microspheres at baseline and during intrabronchial artery challenge with methacholine chloride (MCh; n = 7). Additionally, we measured airway resistance (Raw) during MCh with control or increased perfusion (n = 9). Raw with MCh was significantly greater for high than control flow. Microspheres in histological sections lodged predominantly in the mucosa (60%), and this was not altered by MCh. However, more microspheres lodged in airways >1-mm in diameter during MCh and increased perfusion than MCh and control flow. In airways < or =1 mm in diameter, fewer microspheres lodged during control than increased flow. If the number of microspheres represents regional agonist access to airway smooth muscle, then the differences observed in Raw can be explained by the distribution of agonist. During challenge, there was greater MCh delivery to larger airways during increased flow and less delivery to smaller airways during control flow. The results demonstrate the effects of axial perfusion distribution on Raw.     

Distribution of airway narrowing responses across generations and at branching points,assessed in vitro by anatomical optical coherence tomography

Background

Previous histological and imaging studies have shown the presence of variability in the degree of bronchoconstriction of airways sampled at different locations in the lung (i.e., heterogeneity). Heterogeneity can occur at different airway generations and at branching points in the bronchial tree. Whilst heterogeneity has been detected by previous experimental approaches, its spatial relationship either within or between airways is unknown.

Methods

In this study, distribution of airway narrowing responses across a portion of the porcine bronchial tree was determined in vitro. The portion comprised contiguous airways spanning bronchial generations (#3-11), including the associated side branches. We used a recent optical imaging technique, anatomical optical coherence tomography, to image the bronchial tree in three dimensions. Bronchoconstriction was produced by carbachol administered to either the adventitial or luminal surface of the airway. Luminal cross sectional area was measured before and at different time points after constriction to carbachol and airway narrowing calculated from the percent decrease in luminal cross sectional area.

Results

When administered to the adventitial surface, the degree of airway narrowing was progressively increased from proximal to distal generations (r = 0.80 to 0.98, P < 0.05 to 0.001). This ''serial heterogeneity'' was also apparent when carbachol was administered via the lumen, though it was less pronounced. In contrast, airway narrowing was not different at side branches, and was uniform both in the parent and daughter airways.

Conclusions

Our findings demonstrate that the bronchial tree expresses intrinsic serial heterogeneity, such that narrowing increases from proximal to distal airways, a relationship that is influenced by the route of drug administration but not by structural variations accompanying branching sites.     

A novel bronchial ring bioassay for the evaluation of small airway smooth muscle function in mice

Advances in our understanding of murine airway physiology have been hindered by the lack of suitable, ex vivo, small airway bioassay systems. In this study, we introduce a novel small murine airway bioassay system that permits the physiological and pharmacological study of intrapulmonary bronchial smooth muscle via a bronchial ring (BR) preparation utilizing BR segments as small as 200 microm in diameter. Using this ex vivo BR bioassay, we characterized small airway smooth muscle contraction and relaxation in the presence and absence of bronchial epithelium. In control BRs, the application of mechanical stretch is followed by spontaneous bronchial smooth muscle relaxation. BRs pretreated with methacholine (MCh) partially attenuate this stretch-induced relaxation by as much as 42% compared with control. MCh elicited a dose-dependent bronchial constriction with a maximal tension (E(max)) of 8.7 +/- 0.2 mN at an EC(50) of 0.33 +/- 0.02 microM. In the presence of nifedipine, ryanodine, 2-aminoethoxydiphenyl borate, and SKF-96365, E(max) to MCh was significantly reduced. In epithelium-denuded BRs, MCh-induced contraction was significantly enhanced to 11.4 +/- 1.0 mN with an EC(50) of 0.16 +/- 0.04 microM (P < 0.01). Substance P relaxed MCh-precontracted BR by 62.1%; however, this bronchial relaxation effect was completely lost in epithelium-denuded BRs. Papaverine virtually abolished MCh-induced constriction in both epithelium-intact and epithelium-denuded bronchial smooth muscle. In conclusion, this study introduces a novel murine small airway BR bioassay that allows for the physiological study of smooth muscle airway contractile responses that may aid in our understanding of the pathophysiology of asthma.     

Reduction in airway hyperresponsiveness to methacholine by the application of RF energy in dogs.

We delivered controlled radio frequency energy to the airways of anesthetized, ventilated dogs to examine the effect of this treatment on reducing airway narrowing caused by a known airway constrictor. The airways of 11 dogs were treated with a specially designed bronchial catheter in three of four lung regions. Treatments in each of the three treated lung regions were controlled to a different temperature (55, 65, and 75 degrees C); the untreated lung region served as a control. We measured airway responsiveness to local methacholine chloride (MCh) challenge before and after treatment and examined posttreatment histology to 3 yr. Treatments controlled to 65 degrees C as well as 75 degrees C persistently and significantly reduced airway responsiveness to local MCh challenge (P < or = 0.022). Airway responsiveness (mean percent decrease in airway diameter after MCh challenge) averaged from 6 mo to 3 yr posttreatment was 79 +/- 2.2% in control airways vs. 39 +/- 2.6% (P < or = 0.001) for airways treated at 65 degrees C, and 26 +/- 2.7% (P < or = 0.001) for airways treated at 75 degrees C. Treatment effects were confined to the airway wall and the immediate peribronchial region on histological examination. Airway responsiveness to local MCh challenge was inversely correlated to the extent of altered airway smooth muscle observed in histology (r = -0.54, P < 0.001). We conclude that the temperature-controlled application of radio frequency energy to the airways can reduce airway responsiveness to MCh for at least 3 yr in dogs by reducing airway smooth muscle contractility.     

Differential effects of furosemide on porcine bronchial arterial and airway smooth muscle.

Furosemide attenuates airway obstruction in asthmatic subjects when administered as an aerosol pretreatment. This protective effect of furosemide could be related to relaxation of bronchial smooth muscle or to increased bronchial blood flow. To determine whether furosemide dilates bronchial smooth muscle, isometric contractile responses in distal bronchi from young pigs were studied. In bronchial smooth muscle rings that were precontracted with 10(-5) M acetylcholine, significant relaxation occurred with 10(-8) to 3 x 10(-6) M isoproterenol but not with 10(-8) to 10(-3) M furosemide. In contrast, bronchial arteries that were precontracted with either 10(-4) M norepinephrine or 10(-8) M vasopressin significantly relaxed in response to 10(-4) to 3 x 10(-3) M and 10(-3) to 3 x 10(-3) M furosemide, respectively. We conclude that furosemide, under the described experimental conditions, relaxes airway vascular smooth muscle but not bronchial smooth muscle. These results are consistent with previous suggestions that inhaled furosemide increases blood flow to airway tissues (Gilbert IA, Lenner KA, Nelson JA, Wolin AD, and Fouke JM. J Appl Physiol 76: 409-415, 1994).     

Differences in airway structure in immature and mature rabbits.

Our laboratory has previously demonstrated that maximal bronchoconstriction produces a greater degree of airway narrowing in immature than in mature rabbit lungs (33). To determine whether these maturational differences could be related to airway structure, we compared the fraction of the airway wall occupied by airway smooth muscle (ASM) and cartilage, the proportion of wall area internal to ASM, and the number of alveolar attachments to the airways, from mature and immature (6-mo- and 4-wk-old, respectively) rabbit lungs that were formalin fixed at total lung capacity. The results demonstrate that the airway walls of immature rabbits had a greater percentage of smooth muscle, a lower percentage of cartilage, and fewer alveolar attachments compared with mature rabbit airways; however, we did not find maturational differences in the airway wall thickness relative to airway size. We conclude that structural differences in the airway wall may contribute to the greater airway narrowing observed in immature rabbits during bronchoconstriction.     

In vivo loads on airway smooth muscle: the role of noncontractile airway structures.

The degree of airway smooth muscle contraction and shortening that occurs in vivo is modified by many factors, including those that influence the degree of muscle activation, the resting muscle length, and the loads against which the muscle contracts. Canine trachealis muscle will shorten up to 70% of starting length from optimal length in vitro but will only shorten by around 30% in vivo. This limitation of shortening may be a result of the muscle shortening against an elastic load such as could be applied by tracheal cartilage. Limitation of airway smooth muscle shortening in smaller airways may be the result of contraction against an elastic load, such as could be applied by lung parenchymal recoil. Measurement of the elastic loads applied by the tracheal cartilage to the trachealis muscle and by lung parenchymal recoil to smooth muscle of smaller airways were performed in canine preparations. In both experiments the calculated elastic loads applied by the cartilage and the parenchymal recoil explained in part the limitation of maximal active shortening and airway narrowing observed. We conclude that the elastic loads provided by surrounding structures are important in determining the degree of airway smooth muscle shortening and the resultant airway narrowing.     

Computational model of airway narrowing: mature vs. immature rabbit.

Immature rabbits have greater maximal airway narrowing and greater maximal fold increases in airway resistance during bronchoconstriction than mature animals. We have previously demonstrated that excised immature rabbit lungs have more distensible airways, a lower shear modulus, and structural differences in the relative composition and thickness of anatomically similar airways. In the present study, we incorporated anatomic and physiological data for mature and immature rabbits into a computational model of airway narrowing. We then investigated the relative importance of maturational differences in these factors as determinants of the greater airway narrowing that occurs in the immature animal. The immature model demonstrated greater sensitivity to agonist, as well as a greater maximal fold increase in airway resistance. Exchanging values for airway compliance between the mature and immature models resulted in the mature model exhibiting a greater maximal airway response than the immature model. In contrast, exchanging the shear moduli or the composition of the airway wall relative to the airway size produced relatively small changes in airway reactivity. Our results strongly suggest that the mechanical properties of the airway, i.e., greater compliance of the immature airway, can be an important factor contributing to the greater airway narrowing of the immature animal.     

Effects of airway pressure on bronchial blood flow

We studied the effects of increased airway pressure caused by increasing levels of positive end-expiratory pressure (PEEP) on bronchial arterial pressure-flow relationships. In eight alpha-chloralose-anesthetized mechanically ventilated sheep (23-27 kg), the common bronchial artery, the bronchial branch of the bronchoesophageal artery, was cannulated and perfused with a pump. The control bronchial blood flow (avg 12 +/- 1 ml/min or 0.48 ml X min-1 X kg-1) was set to maintain mean bronchial arterial pressure equal to systemic blood pressure. Pressure-flow curves of the bronchial circulation were measured by making step changes in bronchial blood flow, and changes in these curves were analyzed with measurements of the pressure at zero flow and the slope of the linearized curve. The zero-flow pressure represents the effective downstream pressure, and the slope represents the resistance through the bronchial vasculature. At a constant bronchial arterial pressure of 100 mmHg, an 8 mmHg increase in mean airway pressure caused a 40% reduction in bronchial blood flow. Under constant flow conditions, increases in mean airway pressure with the application of PEEP caused substantial increases in bronchial arterial pressure, averaging 4.6 mmHg for every millimeters of mercury increase in mean airway pressure. However, bronchial arterial pressure at zero flow increased approximately one-for-one with increases in mean airway pressure. Thus the acute sensitivity of the bronchial artery to changes in mean airway pressure results primarily from changes in bronchovascular resistance and not downstream pressure.     

Parenchymal tethering, airway wall stiffness, and the dynamics of bronchoconstriction.

We do not yet have a good quantitative understanding of how the force-velocity properties of airway smooth muscle interact with the opposing loads of parenchymal tethering and airway wall stiffness to produce the dynamics of bronchoconstriction. We therefore developed a two-dimensional computational model of a dynamically narrowing airway embedded in uniformly elastic lung parenchyma and compared the predictions of the model to published measurements of airway resistance made in rats and rabbits during the development of bronchoconstriction following a bolus injection of methacholine. The model accurately reproduced the experimental time-courses of airway resistance as a function of both lung inflation pressure and tidal volume. The model also showed that the stiffness of the airway wall is similar in rats and rabbits, and significantly greater than that of the lung parenchyma. Our results indicate that the main features of the dynamical nature of bronchoconstriction in vivo can be understood in terms of the classic Hill force-velocity relationship operating against elastic loads provided by the surrounding lung parenchyma and an airway wall that is stiffer than the parenchyma.     

Lung parenchymal shear modulus, airway wall remodeling, and bronchial hyperresponsiveness

Lambert, Rodney K., and Peter D. Paré. Lungparenchymal shear modulus, airway wall remodeling, and bronchialhyperresponsiveness. J. Appl. Physiol.83(1): 140-147, 1997.When airways narrow, either through theaction of smooth muscle shortening or during forced expiration, thelung parenchyma is locally distorted and provides an increasedperibronchial stress that resists the narrowing. Although thisinterdependence has been well studied, the quantitative significance ofairway remodeling to interdependence has not been elucidated. We haveused an improved computational model of the bronchial response tosmooth muscle agonists to investigate the relationships between airwaynarrowing (as indicated by airway resistance), parenchymal shearmodulus, adventitial thickening, and inner wall thickening at lungrecoil pressures of 4, 5, and 8 cmH₂O. We have found that, at lowrecoil pressures, decreases in parenchymal shear modulus have asignificant effect that is comparable to that of moderate thickening ofthe airway wall. At higher lung recoil pressures, the effect isnegligible.

     

A model of the mechanics of airway narrowing

To examine the interaction between airway smooth muscle shortening and airway wall thickening on changes in pulmonary resistance, we have developed a model of the tracheobronchial tree that allows simulation of the mechanisms involved in airway narrowing. The model is based on the symmetrical dichotomous branching tracheobronchial tree as described by Weibel and uses fluid dynamic equations proposed by Pedley et al. to calculate inspiratory resistance during quiet tidal breathing. To allow for changes in lung volume, we used the airway pressure-area curves developed by Lambert et al. The model is easily implemented with a spreadsheet and personal computer that allows calculation of total and regional pulmonary resistance. At each airway generation in the model, provision is made for airway wall thickness, the maximal airway smooth muscle shortening achievable, and an S-shaped dose-response relationship to describe smooth muscle shortening. To test the validity of the model, we compared pressure-flow curves generated with the model with measurements of pulmonary resistance while normal subjects breathed air and 20% O2-80% He at a variety of lung volumes. By simulating progressive airway smooth muscle shortening, realistic pulmonary resistance vs. dose-response curves were produced. We conclude that this model provides realistic estimates of pulmonary resistance and shows potential for examining the various mechanisms that could produce excessive airway narrowing in disease.     

Complex airway behavior and paradoxical responses to bronchoprovocation.

Heterogeneity of airway constriction and regional ventilation in asthma are commonly studied under the paradigm that each airway's response is independent from other airways. However, some paradoxical effects and contradictions in recent experimental and theoretical findings suggest that considering interactions among serial and parallel airways may be necessary. To examine airway behavior in a bronchial tree with 12 generations, we used an integrative model of bronchoconstriction, including for each airway the effects of pressure, tethering forces, and smooth muscle forces modulated by tidal stretching during breathing. We introduced a relative smooth muscle activation factor (T(r)) to simulate increasing and decreasing levels of activation. At low levels of T(r), the model exhibited uniform ventilation and homogeneous airway narrowing. But as T(r) reached a critical level, the airway behavior suddenly changed to a dual response with a combination of constriction and dilation. Ventilation decreased dramatically in a group of terminal units but increased in the rest. A local increase of T(r) in a single central airway resulted in full closure, while no central airway closed under global elevation of T(r). Lung volume affected the response to both local and global stimulation. Compared with imaging data for local and global stimuli, as well as for the time course of airway lumen caliber during bronchoconstriction recovery, the model predictions were similar. The results illustrate the relevance of dynamic interactions among serial and parallel pathways in airway interdependence, which may be critical for the understanding of pathological conditions in asthma.     

Angiogenesis and Vascular Remodeling in Chronic Airway Diseases

Asthma and chronic obstructive pulmonary disease remain a global health problem, with increasing morbidity and mortality. Despite differences in the causal agents, both diseases exhibit various degrees of inflammatory changes, structural alterations of the airways leading to airflow limitation. The existence of transient disease phenotypes which overlap both diseases and which progressively decline the lung function has complicated the search for an effective therapy. Important characteristics of chronic airway diseases include airway and vascular remodeling, of which the molecular mechanisms are complex and poorly understood. Recently, we and others have shown that airway smooth muscle (ASM) cells are not only structural and contractile components of airways, rather they bear capabilities of producing large number of pro-inflammatory and mitogenic factors. Increase in size and number of blood vessels both inside and outside the smooth muscle layer as well as hyperemia of bronchial vasculature are contributing factors in airway wall remodeling in patients with chronic airway diseases, proposing for the ongoing mechanisms like angiogenesis and vascular dilatation. We believe that vascular changes directly add to the airway narrowing and hyper-responsiveness by exudation and transudation of proinflammatory mediators, cytokines and growth factors; facilitating trafficking of inflammatory cells; causing oedema of the airway wall and promoting ASM accumulation. One of the key regulators of angiogenesis, vascular endothelial growth factor in concerted action with other endothelial mitogens play pivotal role in regulating bronchial angiogenesis. In this review article we address recent advances in pulmonary angiogenesis and remodelling that contribute in the pathogenesis of chronic airway diseases.