Connecting incremental shear modulus and Poisson's ratio of lung tissue with morphology and rheology of microstructure

The width and curvature of the collagen and elastin fiber bundles in the human pulmonary interalveolar septa and alveolar mouths are measured. The data, together with the known mechanical properties of collagen and elastin fibers, are used to derive the incremental elastic moduli of the lung tissue. The constitutive equation for small incremental stress and strain superposed on a homeostatic inflated lung is linear and isotropic, and characterized by two material constants.     

Mechanical interactions between collagen and proteoglycans: implications for the stability of lung tissue.

Collagen and elastin are thought to dominate the elasticity of the connective tissue including lung parenchyma. The glycosaminoglycans on the proteoglycans may also play a role because osmolarity of interstitial fluid can alter the repulsive forces on the negatively charged glycosaminoglycans, allowing them to collapse or inflate, which can affect the stretching and folding pattern of the fibers. Hence, we hypothesized that the elasticity of lung tissue arises primarily from 1) the topology of the collagen-elastin network and 2) the mechanical interaction between proteoglycans and fibers. We measured the quasi-static, uniaxial stress-strain curves of lung tissue sheets in hypotonic, normal, and hypertonic solutions. We found that the stress-strain curve was sensitive to osmolarity, but this sensitivity decreased after proteoglycan digestion. Images of immunofluorescently labeled collagen networks showed that the fibers follow the alveolar walls that form a hexagonal-like structure. Despite the large heterogeneity, the aspect ratio of the hexagons at 30% uniaxial strain increased linearly with osmolarity. We developed a two-dimensional hexagonal network model of the alveolar structure incorporating the mechanical properties of the collagen-elastin fibers and their interaction with proteoglycans. The model accounted for the stress-strain curves observed under all experimental conditions. The model also predicted how aspect ratio changed with osmolarity and strain, which allowed us to estimate the Young's modulus of a single alveolar wall and a collagen fiber. We therefore identify a novel and important role for the proteoglycans: they stabilize the collagen-elastin network of connective tissues and contribute to lung elasticity and alveolar stability at low to medium lung volumes.     

Spatial distribution of collagen and elastin fibers in the lungs

Surface tension forces acting on the thin-wall alveolar septa and the collagen-elastin fiber network are major factors in lung parenchymal micromechanics. Quantitative serial section analysis and morphometric evaluations of planar sections were used to determine the spatial location of collagen and elastin fibers in Sprague-Dawley rat and normal human lung samples. A large concentration of connective tissue fibers was located in the alveolar duct wall in both species. For rats, the tissue densities of collagen and elastin fibers located within 10 microns of an alveolar duct were 13 and 9%, respectively. In human lung samples, the tissue densities of collagen and elastin fibers within 20 microns of an alveolar duct were 18 and 16%, respectively. In both species, bands of elastin fibers formed a continuous ring around each alveolar mouth. In human lungs, elastin fibers were found to penetrate significantly deeper into alveolar septal walls than they did in rat lungs. The concentration of connective tissue elements in the alveolar duct walls of both species is consistent with their proposed roles as the principal load-bearing elements of the lung parenchyma.     

Arterial Extracellular Matrix: A Mechanobiological Study of the Contributions and Interactions of Elastin and Collagen

The complex network structure of elastin and collagen extracellular matrix (ECM) forms the primary load bearing components in the arterial wall. The structural and mechanobiological interactions between elastin and collagen are important for properly functioning arteries. Here, we examined the elastin and collagen organization, realignment, and recruitment by coupling mechanical loading and multiphoton imaging. Two-photon excitation fluorescence and second harmonic generation methods were performed with a multiphoton video-rate microscope to capture real time changes to the elastin and collagen structure during biaxial deformation. Enzymatic removal of elastin was performed to assess the structural changes of the remaining collagen structure. Quantitative analysis of the structural changes to elastin and collagen was made using a combination of two-dimensional fast Fourier transform and fractal analysis, which allows for a more complete understanding of structural changes. Our study provides new quantitative evidence, to our knowledge on the sequential engagement of different arterial ECM components in response to mechanical loading. The adventitial collagen exists as large wavy bundles of fibers that exhibit fiber engagement after 20% strain. The medial collagen is engaged throughout the stretching process, and prominent elastic fiber engagement is observed up to 20% strain after which the engagement plateaus. The fiber orientation distribution functions show remarkably different changes in the ECM structure in response to mechanical loading. The medial collagen shows an evident preferred circumferential distribution, however the fiber families of adventitial collagen are obscured by their waviness at no or low mechanical strains. Collagen fibers in both layers exhibit significant realignment in response to unequal biaxial loading. The elastic fibers are much more uniformly distributed and remained relatively unchanged due to loading. Removal of elastin produces similar structural changes in collagen as mechanical loading. Our study suggests that the elastic fibers are under tension and impart an intrinsic compressive stress on the collagen.     

Arterial Extracellular Matrix: A Mechanobiological Study of the Contributions and Interactions of Elastin and Collagen

The complex network structure of elastin and collagen extracellular matrix (ECM) forms the primary load bearing components in the arterial wall. The structural and mechanobiological interactions between elastin and collagen are important for properly functioning arteries. Here, we examined the elastin and collagen organization, realignment, and recruitment by coupling mechanical loading and multiphoton imaging. Two-photon excitation fluorescence and second harmonic generation methods were performed with a multiphoton video-rate microscope to capture real time changes to the elastin and collagen structure during biaxial deformation. Enzymatic removal of elastin was performed to assess the structural changes of the remaining collagen structure. Quantitative analysis of the structural changes to elastin and collagen was made using a combination of two-dimensional fast Fourier transform and fractal analysis, which allows for a more complete understanding of structural changes. Our study provides new quantitative evidence, to our knowledge on the sequential engagement of different arterial ECM components in response to mechanical loading. The adventitial collagen exists as large wavy bundles of fibers that exhibit fiber engagement after 20% strain. The medial collagen is engaged throughout the stretching process, and prominent elastic fiber engagement is observed up to 20% strain after which the engagement plateaus. The fiber orientation distribution functions show remarkably different changes in the ECM structure in response to mechanical loading. The medial collagen shows an evident preferred circumferential distribution, however the fiber families of adventitial collagen are obscured by their waviness at no or low mechanical strains. Collagen fibers in both layers exhibit significant realignment in response to unequal biaxial loading. The elastic fibers are much more uniformly distributed and remained relatively unchanged due to loading. Removal of elastin produces similar structural changes in collagen as mechanical loading. Our study suggests that the elastic fibers are under tension and impart an intrinsic compressive stress on the collagen.     

Mechanical forces regulate elastase activity and binding site availability in lung elastin

Many fundamental cellular and extracellular processes in the body are mediated by enzymes. At the single molecule level, enzyme activity is influenced by mechanical forces. However, the effects of mechanical forces on the kinetics of enzymatic reactions in complex tissues with intact extracellular matrix (ECM) have not been identified. Here we report that physiologically relevant macroscopic mechanical forces modify enzyme activity at the molecular level in the ECM of the lung parenchyma. Porcine pancreatic elastase (PPE), which binds to and digests elastin, was fluorescently conjugated (f-PPE) and fluorescent recovery after photobleach was used to evaluate the binding kinetics of f-PPE in the alveolar walls of normal mouse lungs. Fluorescent recovery after photobleach indicated that the dissociation rate constant (k_off) for f-PPE was significantly larger in stretched than in relaxed alveolar walls with a linear relation between k_off and macroscopic strain. Using a network model of the parenchyma, a linear relation was also found between k_off and microscopic strain on elastin fibers. Further, the binding pattern of f-PPE suggested that binding sites on elastin unfold with strain. The increased overall reaction rate also resulted in stronger structural breakdown at the level of alveolar walls, as well as accelerated decay of stiffness and decreased failure stress of the ECM at the macroscopic scale. These results suggest an important role for the coupling between mechanical forces and enzyme activity in ECM breakdown and remodeling in development, and during diseases such as pulmonary emphysema or vascular aneurysm. Our findings may also have broader implications because in vivo, enzyme activity in nearly all cellular and extracellular processes takes place in the presence of mechanical forces.     

Impact of microvascular circulation on peripheral lung stability

The involvement of pulmonary circulation in the mechanical properties was studied in isolated rat lungs. Pulmonary input impedance (ZL) was measured at a mean transpulmonary pressure (Ptpmean) of 2 cmH2O before and after physiological perfusion with either blood or albumin. In these lungs and in a group of unperfused lungs, ZL was also measured at Ptpmean values between 1 and 8 cmH2O. Airway resistance (Raw) and parenchymal damping (G) and elastance (H) were estimated from ZL. End-expiratory lung volume (EELV) was measured by immersion before and after blood perfusion. The orientation of the elastin fibers relative to the basal membrane was assessed in additional unperfused and blood-perfused lungs. Pressurization of the pulmonary capillaries significantly decreased H by 31.5 +/- 3.7% and 18.7 +/- 2.7% for blood and albumin, respectively. Perfusion had no effect on Raw but markedly altered the Ptpmean dependences of G and H < 4 cmH2O, with significantly lower values than in the unperfused lungs. At a Ptpmean of 2 cmH2O, EELV increased by 31 +/- 11% (P = 0.01) following pressurization of the capillaries, and the elastin fibers became more parallel to the basal membrane. Because the organization of elastin fibers results in smaller H values of the individual alveolus, the higher H in the unperfused lungs is probably due to a partial alveolar collapse leading to a loss in lung volume. We conclude that the physiological pressure in the pulmonary capillaries is an important mechanical factor in the maintenance of the stability of the alveolar architecture.     

Structural changes in elastic fibers after pancreatic elastase administration in hamsters.

Ultrastructural changes in lung parenchymal elastic fibers were studied morphometrically 1, 4, and 12 wk after a single 12-unit dose of pancreatic elastase and in a saline-instilled control group. The mean linear intercept of the parenchymal air spaces was increased in the 1-, 4-, and 12-wk post-elastase instillation groups compared with age-matched controls. The volume of alveolar connective tissue fibers predominantly composed of elastin (elastic fibers) was decreased by 35% 1 wk after the instillation of elastase but returned to control levels by 4 wk. Although the total volume of elastic fibers was normal 12 wk after instillation of elastase, the volume of elastic fibers in alveolar entrance rings remained significantly reduced. In serial sections of elastic fibers, numerous gaps or separations in the normally continuous band of elastic fibers that encircle each alveolus were identified 1 wk after elastase instillation. There were 169 +/- 8 (SE), 62 +/- 32, and 12 +/- 6 gaps per millimeter of alveolar entrance ring circumference at 1, 4, and 12 wk, respectively, in the elastase-treated groups. The number of gaps at 12 wk was equivalent to two gaps or discontinuities in the elastic fibers of every alveolar entrance ring. No gaps or separations in elastic fibers were detected at 1, 4, or 12 wk in the control groups. These defects occur in concordance with the progression of air space enlargement and presumably contribute to the progression of air space enlargement that occurs after the elastin content of the tissue has returned to normal.     

A fiber-based constitutive model predicts changes in amount and organization of matrix proteins with development and disease in the mouse aorta

Decreased elastin in mice (Eln+/?) yields a functioning vascular system with elevated blood pressure and increased arterial stiffness that is morphologically distinct from wild-type mice (WT). Yet, function is retained enough that there is no appreciable effect on life span and some mechanical properties are maintained constant. It is not understood how the mouse modifies the normal developmental process to produce a functioning vascular system despite a deficiency in elastin. To quantify changes in mechanical properties, we have applied a fiber-based constitutive model to mechanical data from the ascending aorta during postnatal development of WT and Eln+/? mice. Results indicate that the fiber-based constitutive model is capable of distinguishing elastin amounts and identifying trends during development. We observe an increase in predicted circumferential stress contribution from elastin with age, which correlates with increased elastin amounts from protein quantification data. The model also predicts changes in the unloaded collagen fiber orientation with age, which must be verified in future work. In Eln+/? mice, elastin amounts are decreased at each age, along with the predicted circumferential stress contribution of elastin. Collagen amounts in Eln+/? aorta are comparable to WT, but the predicted circumferential stress contribution of collagen is increased. This may be due to altered organization or structure of the collagen fibers. Relating quantifiable changes in arterial mechanics with changes in extracellular matrix (ECM) protein amounts will help in understanding developmental remodeling and in producing treatments for human diseases affecting ECM proteins.     

Elastin protein levels are a vital modifier affecting normal lung development and susceptibility to emphysema

Cigarette smoking is the strongest risk factor for emphysema. However, sensitivity to cigarette smoke-induced emphysema is highly variable, and numerous genetic and environmental factors are thought to mitigate lung response to injury. We report that the quantity of functional elastin in the lung is an important modifier of both lung development and response to injury. In mice with low levels of elastin, lung development is adversely affected, and mice manifest with congenital emphysema. Animals with intermediate elastin levels exhibit normal alveolar structure but develop worse emphysema than normal mice following cigarette smoke exposure. Mechanical testing demonstrates that lungs with low levels of elastin experience greater tissue strains for any given tissue stress compared with wild-type lungs, implying that force-mediated propagation of lung injury through alveolar wall failure may worsen the emphysema after an initial enzymatic insult. Our findings suggest that quantitative deficiencies in elastin predispose to smoke-induce emphysema in animal models and suggest that humans with altered levels of functional elastin could have relatively normal lung function while being more susceptible to smoke-induced lung injury.     

The role of elastin in the mechanical properties of skin

The elastin fibers of rat skin samples were degraded by the use of a purified preparation of elastase to which soybean inhibitor was added, preventing the collagenolytic activity of the elastase on collagen. Control experiments ascertained degradation of elastin and no effect on collagen. The mechanical properties of the skin samples were studied before and after the enzymatic treatment and differences ascribed to the degraded elastin fibers. Elastin plays a role in the mechanical behaviour of rat skin at small stress values and small deformations. Especially, the elastin fibers are responsible for the recoiling mechanism after a stress or deformation has been applied.     

Stereoscopic particle image velocimetry analysis of healthy and emphysemic alveolar sac models

Emphysema is a progressive lung disease that involves permanent destruction of the alveolar walls. Fluid mechanics in the pulmonary region and how they are altered with the presence of emphysema are not well understood. Much of our understanding of the flow fields occurring in the healthy pulmonary region is based on idealized geometries, and little attention has been paid to emphysemic geometries. The goal of this research was to utilize actual replica lung geometries to gain a better understanding of the mechanisms that govern fluid motion and particle transport in the most distal regions of the lung and to compare the differences that exist between healthy and emphysematous lungs. Excised human healthy and emphysemic lungs were cast, scanned, graphically reconstructed, and used to fabricate clear, hollow, compliant models. Three dimensional flow fields were obtained experimentally using stereoscopic particle image velocimetry techniques for healthy and emphysematic breathing conditions. Measured alveolar velocities ranged over two orders of magnitude from the duct entrance to the wall in both models. Recirculating flow was not found in either the healthy or the emphysematic model, while the average flow rate was three times larger in emphysema as compared to healthy. Diffusion dominated particle flow, which is characteristic in the pulmonary region of the healthy lung, was not seen for emphysema, except for very small particle sizes. Flow speeds dissipated quickly in the healthy lung (60% reduction in 0.25 mm) but not in the emphysematic lung (only 8% reduction 0.25 mm). Alveolar ventilation per unit volume was 30% smaller in emphysema compared to healthy. Destruction of the alveolar walls in emphysema leads to significant differences in flow fields between the healthy and emphysemic lung. Models based on replica geometry provide a useful means to quantify these differences and could ultimately improve our understanding of disease progression.     

Stress failure in pulmonary capillaries

In the mammalian lung, alveolar gas and blood are separated by an extremely thin membrane, despite the fact that mechanical failure could be catastrophic for gas exchange. We raised the pulmonary capillary pressure in anesthetized rabbits until stress failure occurred. At capillary transmural pressures greater than or equal to 40 mmHg, disruption of the capillary endothelium and alveolar epithelium was seen in some locations. The three principal forces acting on the capillary wall were analyzed. 1) Circumferential wall tension caused by the transmural pressure. This is approximately 25 dyn/cm (25 mN/m) at failure where the radius of curvature of the capillary is 5 microns. This tension is small, being comparable with the tension in the alveolar wall associated with lung elastic recoil. 2) Surface tension of the alveolar lining layer. This contributes support to the capillaries that bulge into the alveolar spaces at these high pressures. When protein leakage into the alveolar spaces occurs because of stress failure, the increase in surface tension caused by surfactant inhibition could be a powerful force preventing further failure. 3) Tension of the tissue elements in the alveolar wall associated with lung inflation. This may be negligible at normal lung volumes but considerable at high volumes. Whereas circumferential wall tension is low, capillary wall stress at failure is very high at approximately 8 x 10(5) dyn/cm2 (8 x 10(4) N/m2) where the thickness is only 0.3 microns. This is approximately the same as the wall stress of the normal aorta, which is predominantly composed of collagen and elastin. The strength of the thin part of the capillary wall is probably attributable to the collagen IV of the basement membranes. The safety factor is apparently small when the capillary pressure is raised during heavy exercise. Stress failure causes increased permeability with protein leakage, or frank hemorrhage, and probably has a role in several types of lung disease.     

Role of elastin anisotropy in structural strain energy functions of arterial tissue

The vascular wall exhibits nonlinear anisotropic mechanical properties. The identification of a strain energy function (SEF) is the preferred method to describe its complex nonlinear elastic properties. Earlier constituent-based SEF models, where elastin is modeled as an isotropic material, failed in describing accurately the tissue response to inflation–extension loading. We hypothesized that these shortcomings are partly due to unaccounted anisotropic properties of elastin. We performed inflation–extension tests on common carotid of rabbits before and after enzymatic degradation of elastin and applied constituent-based SEFs, with both an isotropic and an anisotropic elastin part, on the experimental data. We used transmission electron microscopy (TEM) and serial block-face scanning electron microscopy (SBFSEM) to provide direct structural evidence of the assumed anisotropy. In intact arteries, the SEF including anisotropic elastin with one family of fibers in the circumferential direction fitted better the inflation–extension data than the isotropic SEF. This was supported by TEM and SBFSEM imaging, which showed interlamellar elastin fibers in the circumferential direction. In elastin-degraded arteries, both SEFs succeeded equally well in predicting anisotropic wall behavior. In elastase-treated arteries fitted with the anisotropic SEF for elastin, collagen engaged later than in intact arteries. We conclude that constituent-based models with an anisotropic elastin part characterize more accurately the mechanical properties of the arterial wall when compared to models with simply an isotropic elastin. Microstructural imaging based on electron microscopy techniques provided evidence for elastin anisotropy. Finally, the model suggests a later and less abrupt collagen engagement after elastase treatment.     

Effects of elastase on the mechanical and failure properties of engineered elastin-rich matrices.

Pulmonary emphysema and vessel wall aneurysms are diseases characterized by elastolytic damage to elastin fibers that leads to mechanical failure. To model this, neonatal rat aortic smooth muscle cells were cultured, accumulating an extracellular matrix rich in elastin, and mechanical measurements were made before and during enzymatic digestion of elastin. Specifically, the cells in the cultures were killed with sodium azide, the cultures were lifted from the flask, cut into small strips, and fixed to a computer-controlled lever arm and a force transducer. The strips were subjected to a broadband displacement signal to study the dynamic mechanical properties of the samples. Also, quasi-static stress-strain curves were measured. The dynamic data were fit to a linear viscoelastic model to estimate the tissues' loss (G) and storage (H) modulus coefficients, which were evaluated before and during 30 min of elastase treatment, at which point a failure test was performed. G and H decreased significantly to 30% of their baseline values after 30 min. The failure stress of control samples was approximately 15 times higher than that of the digested samples. Understanding the structure-function relationship of elastin networks and the effects of elastolytic injury on their mechanical properties can lead to the elucidation of the mechanism of elastin fiber failure and evaluation of possible treatments to enhance repair in diseases involving elastolytic injury.     

Contribution of elastin and collagen to the inflation response of the pig thoracic aorta: assessing elastin's role in mechanical homeostasis

This study was undertaken to understand elastin's role in the mechanical homeostasis of the arterial wall. The mechanical properties of elastin vary along the aorta, and we hypothesized this maintained a uniform mechanical environment for the elastin, despite regional variation in loading. Elastin's physiological loading was determined by comparing the inflation response of intact and autoclave purified elastin aortas from the proximal and distal thoracic aorta. Elastin's stretch and stress depend on collagen recruitment. Collagen recruitment started in the proximal aorta at systolic pressures (13.3 to 14.6 kPa) and in the distal at sub-diastolic pressures (9.3 to 10.6 kPa). In the proximal aorta collagen did not contribute significantly to the stress or stiffness, indicating that elastin determined the vessel properties. In the distal aorta, the circumferential incremental modulus was 70% higher than in the proximal aorta, half of which (37%) was due to a stiffening of the elastin. Compared to the elastin tissue in the proximal aorta, the distal elastin suffered higher physiological circumferential stretch (29%, P=0.03), circumferential stress (39%, P=0.02), and circumferential stiffness (37%, P=0.006). Elastin's physiological axial stresses were also higher (67%, P=0.003). These findings do not support the hypothesis that the loading on elastin is constant along the aorta as we expected from homeostasis.     

Comparison of amounts of collagen and elastin in pleura and parenchyma of dog lung

Pressure-volume characteristics of the lung have been thought to be due primarily to the properties of the network of alveolar septa. However, Hajji et al. (J. Appl. Physiol.: Respirat . Environ. Exercise Physiol. 47: 175-181, 1979) attributed a substantial role to the visceral pleura. Seeking a structural explanation for this result, we compared the relative amounts of collagen fibrils and elastin fibers in the visceral pleura and alveolar parenchyma using stereological measurements in five canine lobes. We found about one-fifth as much collagen and one-tenth as much elastin in the pleura as in the alveolar parenchyma. This structural result confirms the functional conclusions of Hajji et al. We argue that such a substantial structure is not needed for protection against overinflation but may have to do with stabilization of lobe shape or handling of frictional forces.     

Time course of lung parenchyma remodeling in pulmonary and extrapulmonary acute lung injury.

The aim of this study is to test the hypothesis that the early changes in lung mechanics and the amount of type III collagen fiber do not predict the evolution of lung parenchyma remodeling in pulmonary and extrapulmonary acute lung injury (ALI). For this purpose, we analyzed the time course of lung parenchyma remodeling in murine models of pulmonary and extrapulmonary ALI with similar degrees of mechanical compromise at the early phase of ALI. Lung histology (light and electron microscopy), the amount of elastic and collagen fibers in the alveolar septa, the expression of matrix metalloproteinase-9, and mechanical parameters (lung-resistive and viscoelastic pressures, and static elastance) were analyzed 24 h, 1, 3, and 8 wk after the induction of lung injury. In control (C) pulmonary (p) and extrapulmonary (exp) groups, saline was intratracheally (it; 0.05 ml) instilled and intraperitoneally (ip; 0.5 ml) injected, respectively. In ALIp and ALIexp groups, mice received Escherichia coli lipopolysaccharide (10 microg it and 125 microg ip, respectively). At 24 h, all mechanical and morphometrical parameters, as well as type III collagen fiber content, increased similarly in ALIp and ALIexp groups. In ALIexp, all mechanical and histological data returned to control values at 1 wk. However, in ALIp, static elastance returned to control values at 3 wk, whereas resistive and viscoelastic pressures, as well as type III collagen fibers and elastin, remained elevated until week 8. ALIp showed higher expression of matrix metalloproteinase-9 than ALIexp. In conclusion, insult in pulmonary epithelium yielded fibroelastogenesis, whereas mice with ALI induced by endothelial lesion developed only fibrosis that was repaired early in the course of lung injury. Furthermore, early functional and morphological changes did not predict lung parenchyma remodeling.     

Perfusion dehydration fixes elastin and preserves lung air-space dimensions

We sought a technique to preserve lung tissue for micrography with air-space dimensions unchanged from the fresh state. In our hands, conventional techniques were problematical. Aware of the possibility that distortion might be caused by inadequate mechanical fixation of elastin, we dehydrated the still-inflated lung by intravascular perfusion with graded ethanols. Canine and rabbit lungs so prepared had straighter alveolar septa, greater air-space dimensions, and an improved correlation with light-scattering measurements. Bovine ligamentum nuchae (mostly elastin) was only partially fixed by glutaraldehyde or osmium tetroxide but was effectively stiffened by dehydration. We conclude that perfusion dehydration aids in the faithful preservation of parenchymal configuration, probably by mechanical fixation of elastin.     

Electron microscopic observations of elastic fibres in the lung and aorta of tight-skin and beta-aminopropionitrile-fed mice.

The lung of the tight-skin (TSK) mouse was characterized by enlargement of the air spaces. Elastin in the alveolar walls of the TSK mouse exhibited fragmentation. The aorta of the TSK mouse was characterized by marked hyperplasia of loose connective tissue in the adventitia. Collagen fibres and ruthenium red-positive materials were markedly increased. Microfibrils surrounding elastin in the adventitia of the aorta were not clear in the TSK mouse. In the lung of the beta-aminopropionitrile (BAPN)-fed mouse, enlargement of the alveolar air spaces was not prominent compared with the TSK mouse. Elastic fibres in the alveolar walls did not show the fragmentation observed in the TSK mouse, and microfibrils surrounding elastin were clearly observed. However, elastic laminae in the media of the BAPN-fed mouse aorta were swollen and fragmented. Elastic fibres in the adventitia exhibited a normal appearance and microfibrils surrounding elastin in the adventitia were clearly observed. The results suggest that the mechanism of the connective tissue abnormality in the TSK mouse is different from that of BAPN, which inhibits the activity of lysyl oxidase. The abnormality of elastin and microfibrils surrounding elastin in the TSK mouse probably plays a role in the deformity or degradation of elastic fibres and the structural changes of the lung.