Elucidating the interaction between stretch and stiffness using an agent-based spring network model of progressive pulmonary fibrosis.

Pulmonary fibrosis is a deadly disease that involves the dysregulation of fibroblasts and myofibroblasts, which are mechanosensitive. Previous computational models have succeeded in modeling stiffness-mediated fibroblasts behaviors; however, these models have neglected to consider stretch-mediated behaviors, especially stretch-sensitive channels and the stretch-mediated release of latent TGF-ß. Here, we develop and explore an agent-based model and spring network model hybrid that is capable of recapitulating both stiffness and stretch. Using the model, we evaluate the role of mechanical signaling in homeostasis and disease progression during self-healing and fibrosis, respectively. We develop the model such that there is a fibrotic threshold near which the network tends towards instability and fibrosis or below which the network tends to heal. The healing response is due to the stretch signal, whereas the fibrotic response occurs when the stiffness signal overpowers the stretch signal, creating a positive feedback loop. We also find that by changing the proportional weights of the stretch and stiffness signals, we observe heterogeneity in pathological network structure similar to that seen in human IPF tissue. The system also shows emergent behavior and bifurcations: whether the network will heal or turn fibrotic depends on the initial network organization of the damage, clearly demonstrating structure's pivotal role in healing or fibrosis of the overall network. In summary, these results strongly suggest that the mechanical signaling present in the lungs combined with network effects contribute to both homeostasis and disease progression.

Local fractal dimension of collagen detects increased spatial complexity in fibrosis.

Increase of collagen content and reorganization characterizes fibrosis but quantifying the latter remains challenging. Spatially complex structures are often analyzed via the fractal dimension; however, established methods for calculating this quantity either provide a single dimension for an entire object or a spatially distributed dimension that only considers binary images. These neglect valuable information related to collagen density in images of fibrotic tissue. We sought to develop a fractal analysis that can be applied to 3-dimensional (3D) images of fibrotic tissue. A fractal dimension map for each image was calculated by determining a single fractal dimension for a small area surrounding each image pixel, using fiber thickness as the third dimension. We found that this local fractal dimension increased with age and with progression of fibrosis regardless of collagen content. Our new method of distributed 3D fractal analysis can thus distinguish between changes in collagen content and organization induced by fibrosis.

Crystal ribcage: a platform for probing real-time lung function at cellular resolution.

Understanding the dynamic pathogenesis and treatment response in pulmonary diseases requires probing the lung at cellular resolution in real time. Despite advances in intravital imaging, optical imaging of the lung during active respiration and circulation has remained challenging. Here, we introduce the crystal ribcage: a transparent ribcage that allows multiscale optical imaging of the functioning lung from whole-organ to single-cell level. It enables the modulation of lung biophysics and immunity through intravascular, intrapulmonary, intraparenchymal and optogenetic interventions, and it preserves the three-dimensional architecture, air-liquid interface, cellular diversity and respiratory-circulatory functions of the lung. Utilizing these capabilities on murine models of pulmonary pathologies we probed remodeling of respiratory-circulatory functions at the single-alveolus and capillary levels during disease progression. The crystal ribcage and its broad applications presented here will facilitate further studies of nearly any pulmonary disease as well as lead to the identification of new targets for treatment strategies.

Multiscale stiffness of human emphysematous precision cut lung slices.

Emphysema is a debilitating disease that remodels the lung leading to reduced tissue stiffness. Thus, understanding emphysema progression requires assessing lung stiffness at both the tissue and alveolar scales. Here, we introduce an approach to determine multiscale tissue stiffness and apply it to precision-cut lung slices (PCLS). First, we established a framework for measuring stiffness of thin, disk-like samples. We then designed a device to verify this concept and validated its measuring capabilities using known samples. Next, we compared healthy and emphysematous human PCLS and found that the latter was 50% softer. Through computational network modeling, we discovered that this reduced macroscopic tissue stiffness was due to both microscopic septal wall remodeling and structural deterioration. Lastly, through protein expression profiling, we identified a wide spectrum of enzymes that can drive septal wall remodeling, which, together with mechanical forces, lead to rupture and structural deterioration of the emphysematous lung parenchyma.

An agent-based model of tissue maintenance and self-repair.

We hypothesized that a system that possesses the capacity for ongoing maintenance of its tissues will necessarily also have the capacity to self-heal following a perturbation. We used an agent-based model of tissue maintenance to investigate this idea, and in particular to determine the extent to which the current state of the tissue must influence cell behavior in order for tissue maintenance and self-healing to be stable. We show that a mean level of tissue density is robustly maintained when catabolic agents digest tissue at a rate proportional to local tissue density, but that the spatial heterogeneity of the tissue at homeostasis increases with the rate at which tissue is digested. The rate of self-healing is also increased by increasing either the amount of tissue removed or deposited at each time step by catabolic or anabolic agents, respectively, and by increasing the density of both agent types on the tissue. We also found that tissue maintenance and self-healing are stable with an alternate rule in which cells move preferentially to tissue regions of low density. The most basic form of self-healing can thus be achieved with cells that follow very simple rules of behavior, provided these rules are based in some way on the current state of the local tissue. Straightforward mechanisms can accelerate the rate of self-healing, as might be beneficial to the organism.

Predicting alveolar ventilation heterogeneity in pulmonary fibrosis using a non-uniform polyhedral spring network model.

Pulmonary Fibrosis (PF) is a deadly disease that has limited treatment options and is caused by excessive deposition and cross-linking of collagen leading to stiffening of the lung parenchyma. The link between lung structure and function in PF remains poorly understood, although its spatially heterogeneous nature has important implications for alveolar ventilation. Computational models of lung parenchyma utilize uniform arrays of space-filling shapes to represent individual alveoli, but have inherent anisotropy, whereas actual lung tissue is isotropic on average. We developed a novel Voronoi-based 3D spring network model of the lung parenchyma, the Amorphous Network, that exhibits more 2D and 3D similarity to lung geometry than regular polyhedral networks. In contrast to regular networks that show anisotropic force transmission, the structural randomness in the Amorphous Network dissipates this anisotropy with important implications for mechanotransduction. We then added agents to the network that were allowed to carry out a random walk to mimic the migratory behavior of fibroblasts. To model progressive fibrosis, agents were moved around the network and increased the stiffness of springs along their path. Agents migrated at various path lengths until a certain percentage of the network was stiffened. Alveolar ventilation heterogeneity increased with both percent of the network stiffened, and walk length of the agents, until the percolation threshold was reached. The bulk modulus of the network also increased with both percent of network stiffened and path length. This model thus represents a step forward in the creation of physiologically accurate computational models of lung tissue disease.

Time dependency and unique etiology of barotrauma in COVID-19: A retrospective cohort study with landmark analysis and pathological approach.

BACKGROUND: Barotrauma frequently occurs in coronavirus disease 2019. Previous studies have reported barotrauma to be a mortality-risk factor; however, its time-dependent nature and pathophysiology are not elucidated. To investigate the time-dependent characteristics and the etiology of coronavirus disease 2019-related-barotrauma. METHODS AND FINDINGS: We retrospectively reviewed intubated patients with coronavirus disease 2019 from March 2020 to May 2021. We compared the 90-day survival between the barotrauma and non-barotrauma groups and performed landmark analyses on days 7, 14, 21, and 28. Barotrauma within seven days before the landmark was defined as the exposure. Additionally, we evaluated surgically treated cases of coronavirus disease 2019-related pneumothorax. We included 192 patients. Barotrauma developed in 44 patients (22.9%). The barotrauma group's 90-day survival rate was significantly worse (47.7% vs. 82.4%, p < 0.001). In the 7-day landmark analysis, there was no significant difference (75.0% vs. 75.7%, p = 0.79). Contrastingly, in the 14-, 21-, and 28-day landmark analyses, the barotrauma group's survival rates were significantly worse (14-day: 41.7% vs. 69.1%, p = 0.044; 21-day: 16.7% vs. 62.5%, p = 0.014; 28-day: 20.0% vs. 66.7%, p = 0.018). Pathological examination revealed a subpleural hematoma and pulmonary cyst with heterogenous lung inflammation. CONCLUSIONS: Barotrauma was a poor prognostic factor for coronavirus disease 2019, especially in the late phase. Heterogenous inflammation may be a key finding in its mechanism. Barotrauma is a potentially important sign of lung destruction.

Modeling the influence of gravity and the mechanical properties of elastin and collagen fibers on alveolar and lung pressure-volume curves.

The relationship between pressure (P) and volume (V) in the human lung has been extensively studied. However, the combined effects of gravity and the mechanical properties of elastin and collagen on alveolar and lung P-V curves during breathing are not well understood. Here, we extended a previously established thick-walled spherical model of a single alveolus with wavy collagen fibers during positive pressure inflation. First, we updated the model for negative pressure-driven inflation that allowed incorporation of a gravity-induced pleural pressure gradient to predict how the static alveolar P-V relations vary spatially throughout an upright human lung. Second, by introducing dynamic surface tension and collagen viscoelasticity, we computed the hysteresis loop of the lung P-V curve. The model was tested by comparing its predicted regional ventilation to literature data, which offered insight into the effects of microgravity on ventilation. The model has also produced novel testable predictions for future experiments about the variation of mechanical stresses in the septal walls and the contribution of collagen and elastin fibers to the P-V curve and throughout the lung. The model may help us better understand how mechanical stresses arising from breathing and pleural pressure variations affect regional cellular mechanotransduction in the lung.

Remodeling of the Aged and Emphysematous Lungs: Roles of Microenvironmental Cues.

Aging is a slow process that affects all organs, and the lung is no exception. At the alveolar level, aging increases the airspace size with thicker and stiffer septal walls and straighter and thickened collagen and elastic fibers. This creates a microenvironment that interferes with the ability of cells in the parenchyma to maintain normal homeostasis and respond to injury. These changes also make the lung more susceptible to disease such as emphysema. Emphysema is characterized by slow but progressive remodeling of the deep alveolar regions that leads to airspace enlargement and increased but disorganized elastin and collagen deposition. This remodeling has been attributed to ongoing inflammation that involves inflammatory cells and the cytokines they produce. Cellular senescence, another consequence of aging, weakens the ability of cells to properly respond to injury, something that also occurs in emphysema. These factors conspire to make alveolar walls more prone to mechanical failure, which can set emphysema in motion by driving inflammation through immune stimulation by protein fragments. Both aging and emphysema are influenced by microenvironmental conditions such as local inflammation, chemical makeup, tissue stiffness, and mechanical stresses. Although aging and emphysema are not equivalent, they have the potential to influence each other in synergistic ways; aging sets up the conditions for emphysema to develop, while emphysema may accelerate cellular senescence and thus aging itself. This article focuses on the similarities and differences between the remodeled microenvironment of the aging and emphysematous lung, with special emphasis on the alveolar septal wall. © 2022 American Physiological Society. Compr Physiol 12:3559-3574, 2022.

Harmonic Distortion of Blood Pressure Waveform as a Measure of Arterial Stiffness.

Aging and disease alter the composition and elastic properties of the aortic wall resulting in shape changes in blood pressure waveform (BPW). Here, we propose a new index, harmonic distortion (HD), to characterize BPW and its relationship with other in vitro and in vivo measures. Using a Fourier transform of the BPW, HD is calculated as the ratio of energy above the fundamental frequency to that at the fundamental frequency. Male mice fed either a normal diet (ND) or a high fat, high sucrose (HFHS) diet for 2-10 months were used to study BPWs in diet-induced metabolic syndrome. BPWs were recorded for 20 s hourly for 24 h, using radiotelemetry. Pulse wave velocity (PWV), an in vivo measure of arterial stiffness, was measured in the abdominal aorta via ultrasound sonography. Common carotid arteries were excised from a subset of mice to determine the tangent modulus using biaxial tension-inflation test. Over a 24-h period, both HD and systolic blood pressure (SBP) show a large variability, however HD linearly decreases with increasing SBP. HD is also linearly related to tangent modulus and PWV with slopes significantly different between the two diet groups. Overall, our study suggests that HD is sensitive to changes in blood pressure and arterial stiffness and has a potential to be used as a noninvasive measure of arterial stiffness in aging and disease.

A Personalized Spring Network Representation of Emphysematous Lungs From CT Images.

Emphysema is a progressive disease characterized by irreversible tissue destruction and airspace enlargement, which manifest as low attenuation area (LAA) on CT images. Previous studies have shown that inflammation, protease imbalance, extracellular matrix remodeling and mechanical forces collectively influence the progression of emphysema. Elastic spring network models incorporating force-based mechanical failure have been applied to investigate the pathogenesis and progression of emphysema. However, these models were general without considering the patient-specific information on lung structure available in CT images. The aim of this work was to develop a novel approach that provides an optimal spring network representation of emphysematous lungs based on the apparent density in CT images, allowing the construction of personalized networks. The proposed method takes into account the size and curvature of LAA clusters on the CT images that correspond to a pre-stressed condition of the lung as opposed to a naïve method that excludes the effects of pre-stress. The main findings of this study are that networks constructed by the new method 1) better preserve LAA cluster sizes and their distribution than the naïve method; and 2) predict different course of emphysema progression compared to the naïve method. We conclude that our new method has the potential to predict patient-specific emphysema progression which needs verification using clinical data.

Mechano-inflammatory sensitivity of ACE2: Implications for the regional distribution of SARS-CoV-2 injury in the lung.

The coronavirus disease (COVID-19) caused by SARS-CoV-2 can result in severe injury to the lung. Computed tomography images have revealed that the virus preferentially affects the base of the lung, which experiences larger tidal stretches than the apex. We hypothesize that the expression of both the angiotensin converting enzyme-2 (ACE2) receptor for SARS-CoV-2 and the transmembrane serine protease 2 (TMPRSS2) are sensitive to regional cell stretch in the lung. To test this hypothesis, we stretched precision cut lung slices (PCLS) for 12 h with one of the following protocols: 1) unstretched (US); 2) low-stretch (LS), 5% peak-to-peak area strain mimicking the lung base; or 3) high-stretch (HS), the same peak-to-peak area strain superimposed on 10% static area stretch mimicking the lung apex. PCLS were additionally stretched in cigarette smoke extract (CSE) to mimic an acute inflammatory exposure. The expression of ACE2 was higher whereas that of TMPRSS2 was lower in the control samples following LS than HS. CSE-induced inflammation substantially altered the expression of ACE2 with higher levels following HS than LS. These results suggest that ACE2 and TMPRSS2 expression in lung cells is mechanosensitive, which could have implications for the spatial distribution of COVID-19-mediated lung injury and the increased risk for more severe disease in active smokers and patients with COPD.

Inflation instability in the lung: an analytical model of a thick-walled alveolus with wavy fibres under large deformations.

Inflation of hollow elastic structures can become unstable and exhibit a runaway phenomenon if the tension in their walls does not rise rapidly enough with increasing volume. Biological systems avoid such inflation instability for reasons that remain poorly understood. This is best exemplified by the lung, which inflates over its functional volume range without instability. The goal of this study was to determine how the constituents of lung parenchyma determine tissue stresses that protect alveoli from instability-related overdistension during inflation. We present an analytical model of a thick-walled alveolus composed of wavy elastic fibres, and investigate its pressure-volume behaviour under large deformations. Using second-harmonic generation imaging, we found that collagen waviness follows a beta distribution. Using this distribution to fit human pressure-volume curves, we estimated collagen and elastin effective stiffnesses to be 1247 kPa and 18.3 kPa, respectively. Furthermore, we demonstrate that linearly elastic but wavy collagen fibres are sufficient to achieve inflation stability within the physiological pressure range if the alveolar thickness-to-radius ratio is greater than 0.05. Our model thus identifies the constraints on alveolar geometry and collagen waviness required for inflation stability and provides a multiscale link between alveolar pressure and stresses on fibres in healthy and diseased lungs.

Stabilizing breathing pattern using local mechanical vibrations: comparison of deterministic and stochastic stimulations in rodent models of apnea of prematurity.

Mechanical stimulation has been shown to reduce apnea of prematurity (AOP), a major concern in preterm infants. Previous work suggested that the underlying mechanism is stochastic resonance, amplification of a subthreshold signal by stochastic stimulation. We hypothesized that the mechanism behind the reduction of apnea length may not be a solely stochastic phenomenon, and suggest that a purely deterministic, non-random mechanical stimulation could be equally as effective. Mice and rats were anesthetized, tracheostomized, and mechanically ventilated to halt spontaneous breathing. Two miniature motors controlled by a microcontroller were attached around the abdomen. Ventilation was paused, stimulations were applied, and the time to the rodent's first spontaneous breath (T) was measured. Six spectrally different signals were compared to one another and the no-stimulation control in mice. The most successful deterministic stimulation (D) at reducing apnea was then compared to a pseudo-random noise (PRN) signal of comparable amplitude and frequency. CO2%, CO2 stabilization time (Ts), O2 saturation (SpO2%), and T were also measured. D significantly reduced T compared to no stimulation for medium and high amplitudes. PRN also reduced T, without  a difference between D and PRN. Furthermore, both stimulations significantly reduced Ts with no significant differences between the respective stimulations. However, there was no effect of D or PRN on SpO2%. The lack of differences between D and PRN led to an additional series of experiment comparing the same D to a band-limited white noise (WN) signal in young rats. Both D and WN were shown to significantly reduce T, with D showing statistical superiority in reduction of apnea. We further speculate that both deterministic and stochastic mechanical stimulations induce some form of mechanotransduction which is responsible for their efficacy, and our findings suggest that mechanical stimulation may be effective in treating AOP. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13534-021-00203-x.

Percolation of collagen stress in a random network model of the alveolar wall.

Fibrotic diseases are characterized by progressive and often irreversible scarring of connective tissue in various organs, leading to substantial changes in tissue mechanics largely as a result of alterations in collagen structure. This is particularly important in the lung because its bulk modulus is so critical to the volume changes that take place during breathing. Nevertheless, it remains unclear how fibrotic abnormalities in the mechanical properties of pulmonary connective tissue can be linked to the stiffening of its individual collagen fibers. To address this question, we developed a network model of randomly oriented collagen and elastin fibers to represent pulmonary alveolar wall tissue. We show that the stress-strain behavior of this model arises via the interactions of collagen and elastin fiber networks and is critically dependent on the relative fiber stiffnesses of the individual collagen and elastin fibers themselves. We also show that the progression from linear to nonlinear stress-strain behavior of the model is associated with the percolation of stress across the collagen fiber network, but that the location of the percolation threshold is influenced by the waviness of collagen fibers.

Breath Hold Facilitates Targeted Deposition of Aerosolized Droplets in a 3D Printed Bifurcating Airway Tree.

The lungs have long been considered a desired route for drug delivery but, there is still a lack of strategies to rationally target delivery sites especially in the presence of heterogeneous airway disease. Furthermore, no standardized system has been proposed to rapidly test different ventilation strategies and how they alter the overall and regional deposition pattern in the airways. In this study, a 3D printed symmetric bifurcating tree model mimicking part of the human airway tree was developed that can be used to quantify the regional deposition patterns of different delivery methodologies. The model is constructed in a novel way that allows for repeated measurements of regional deposition using reusable parts. During ventilation, nebulized ~3-micron-sized fluid droplets were delivered into the model. Regional delivery, quantified by precision weighing individual airways, was highly reproducible. A successful strategy to control regional deposition was achieved by combining an inspiratory wave form with a "breath hold" pause after each inspiration. Specifically, the second generation of the tree was successfully targeted, and deposition was increased by up to four times in generation 2 when compared to a ventilation without the breath hold (p < 0.0001). Breath hold was also demonstrated to facilitate deposition into blocked regions of the model, which mimic airway closure during an asthma that receive no flow during inhalation. Additionally, visualization experiments demonstrated that in the absence of fluid flow, the deposition of 3-micron water droplets is dominated by gravity, which, to our knowledge, has not been confirmed under standard laboratory conditions.

Modeling Lung Derecruitment in VILI Due to Fluid-Occlusion: The Role of Emergent Behavior.

Ventilator-induced lung injury (VILI) is driven by the processes of volutrauma and atelectrauma, which can act synergistically to compromise the blood-gas barrier. We have postulated that this synergy arises through a rich-get-richer mechanism whereby atelectrauma causes holes to form in the blood-gas barrier while concomitant volutrauma causes susceptible holes to progressively enlarge as VILI worsens. We previously developed an analytical model based on this idea that accurately predicts the progressive increases in lung elastance seen immediately following a recruitment maneuver as VILI progresses over the course of hours. In the present study we extend this model to account for the rate of change of elastance, due to closure of lung units, in the minutes following a recruitment maneuver. We found that the distribution of unit closing velocities throughout the lung can be described by a power law with an exponent of -2 that matches previously published power laws associated with the dynamics of lung recruitment. Our model thus reveals lung collapse as an example of emergent complex behavior and links the dynamics of altered function in the injured lung to structural damage in a way that explains the mechanisms of injury progression arising from the ongoing stresses and strains applied by mechanical ventilation.