Multiscale Mathematical Modeling of the Absorptive and Mucociliary Pathophysiology of Cystic Fibrosis Lung Disease

Airway disease is the primary cause of mortality for the over 70,000 patients with Cystic Fibrosis (CF) worldwide. It is characterized by lung infection, inflammation, and impaired mucociliary clearance (MCC) arising from depletion of the airway surface liquid (ASL) at the organ-scale. Dysfunction in the CF transmembrane conductance regulator protein causes dysregulation in ion and liquid transport alone and via other transport-related proteins. Analysis of cell-system interactions is experimentally complex, however, and motivates the use of mechanistic mathematical models that can also be used to design and optimize treatments for the disease. Tc99m or In111-labeled DTPA (DTPA) are small-molecule radiological probes that allow for observation of paracellular liquid convection and solute transport at cellular and organ scales, respectively. Previous work has shown that DTPA is hyperabsorbed in CF in a manner that strongly correlates with ASL hyperabsorption. The models of this dissertation describe, in part, the mechanisms that underlie this correlation. At the lung-scale, a physiologically motivated pharmacokinetic model was developed to describe the action of hypertonic saline (HS) as an inhaled therapy in CF. This model predicts that MCC is reduced in patients with CF because they have a reduced fraction of functional ciliated airway -- a model parameter -- that is increased via HS-induced airway rehydration. This prediction was verified \textit{in vitro} in human bronchial epithelial (HBE) cultures. A separate, cell-scale model accurately characterizes transcellular liquid transport in HBE cultures using transport parameters that agree well with previously reported values, producing ion flux estimates from the model fit to ASL and DTPA absorption that were similar to known physiological values. It also implicates diminished constitutive Cl¬- secretion in ASL dehydration but suggests that reduced paracellular integrity is the predominant factor leading to hyperabsorption in CF. The cell- and lung-scale models were then used to analyze treatment failure and suggest modifications of a clinical trial, which is the first indication of the utility of airway transport models in designing treatments for patients with CF

A physiologically-motivated compartment-based model of the effect of inhaled hypertonic saline on mucociliary clearance and liquid transport in cystic fibrosis

Background: Cystic Fibrosis (CF) lung disease is characterized by liquid hyperabsorption, airway surface dehydration, and impaired mucociliary clearance (MCC). Herein, we present a compartment-based mathematical model of the airway that extends the resolution of functional imaging data. Methods: Using functional imaging data to inform our model, we developed a system of mechanism-motivated ordinary differential equations to describe the mucociliary clearance and absorption of aerosolized radiolabeled particle and small molecules probes from human subjects with and without CF. We also utilized a novel imaging metric in vitro to gauge the fraction of airway epithelial cells that have functional ciliary activity. Results: This model, and its incorporated kinetic rate parameters, captures the MCC and liquid dynamics of the hyperabsorptive state in CF airways and the mitigation of that state by hypertonic saline treatment. Conclusions: We postulate, based on the model structure and its ability to capture clinical patient data, that patients with CF have regions of airway with diminished MCC function that can be recruited with hypertonic saline treatment. In so doing, this model structure not only makes a case for durable osmotic agents used in lung-region specific treatments, but also may provide a possible clinical endpoint, the fraction of functional ciliated airway

Mucus, mucins, and cystic fibrosis

Cystic fibrosis (CF) is both the most common and most lethal genetic disease in the Caucasian population. CF is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene and is characterized by the accumulation of thick, adherent mucus plaques in multiple organs, of which the lungs, gastrointestinal tract and pancreatic ducts are the most commonly affected. A similar pathogenesis cascade is observed in all of these organs: loss of CFTR function leads to altered ion transport, consisting of decreased chloride and bicarbonate secretion via the CFTR channel and increased sodium absorption via epithelial sodium channel upregulation. Mucosa exposed to changes in ionic concentrations sustain severe pathophysiological consequences. Altered mucus biophysical properties and weakened innate defense mechanisms ensue, furthering the progression of the disease. Mucins, the high-molecular-weight glycoproteins responsible for the viscoelastic properties of the mucus, play a key role in the disease but the actual mechanism of mucus accumulation is still undetermined. Multiple hypotheses regarding the impact of CFTR malfunction on mucus have been proposed and are reviewed here. (a) Dehydration increases mucin monomer entanglement, (b) defective Ca2+ chelation compromises mucin expansion, (c) ionic changes alter mucin interactions, and (d) reactive oxygen species increase mucin crosslinking. Although one biochemical change may dominate, it is likely that all of these mechanisms play some role in the progression of CF disease. This article discusses recent findings on the initial cause(s) of aberrant mucus properties in CF and examines therapeutic approaches aimed at correcting mucus properties

Assessing the Effectiveness of Shah\u27s Innovation Metrics for Measuring Innovative Design within a Virtual Design Space

Epistemic games, such as the virtual engineering internship Nephrotex, allow students to explore creative ways to approaching engineering problems while providing a novel alternative to the direct transmission method of instruction. Within Nephrotex, students choose a polymer, manufacturing process, surfactant, and percentage of carbon nanotube to create a functioning kidney dialysis membrane prototype. The performance of the membrane is measured using cost, flux, blood cell reactivity, marketability, and reliability thresholds given by stakeholders within the fictitious company. Although Nephrotex has been shown to be a valuable educational tool for modeling the product design process, only limited work has been done to investigate whether it is capable of providing an environment that allows students to generate innovative designs. The innovation assessment framework of Shah and colleagues employs four metrics of innovation – novelty, variety, quality, and quantity; novelty is further divided into a priori and a posteriori metrics. This work found that a priori and a posteriori novelty, variety, and quality were applicable metrics of innovation in the epistemic game environment of Nephrotex. Literature ranges for a priori and a posteriori novelty scores aligned with those found in this study. Comparing prior work on Nephrotex that identified innovative student designs based on a proposed literature definition, it was found that the Shah metrics between the innovative and non-innovative groups showed little variation and no statistically significant differences. A t test and a Mann-Whitney U test showed no significant difference between innovative and non-innovative groups with regard to variety or novelty scores; however, these tests did show a significant difference between groups with regards to the quality score. The same results were found when calculating Cohen’s Effect Size – a priori novelty, a posteriori novelty, and variety had a small effect when comparisons were made between the innovative and non-innovative groups while quality had a large effect. The significant difference and large effect in regards to quality however, may be the result of the previous literature definition which employed quality as a measure to define innovation. Results from this study demonstrate that novelty is perhaps the most aligned innovation metric for an epistemic game environment and that both variety and quality can be helpful in understanding the designs generated within these contexts although they may need adjustment based on the application to a constrained design space

Innovative Design within the Context of Virtual Internships: How Can It Be Defined and How is It Related to the Student Design Process?

Definitions of “innovative design” vary among authors and fields of study. This makes it difficult to establish how to determine innovative designs in new design environments, such as in a virtual internship environment. Nephrotex is a virtual internship which encourages players, who assume the role of virtual interns within the game, to fully explore a constrained design space with the goal of producing an optimized dialysis membrane. As a useful starting point for our definition of “innovative design” within this design environment, we referenced the work of Baregheh et al. (2009) and formed the following definition: “A process that not only leads to unique physical or technical product attributes but also adds value beyond existing designs on the market.” We defined uniqueness based on the design occurring infrequently amongst the final products of student design teams. Quality was assessed based on the work of Arastoopour and colleagues (2014), taking technical and economic performance into consideration to determine how well the design was able to meet the Nephrotex internal consultant requirements for the design. Using this definition, we sought to answer the following research question: How does the design process differ for a team that generates an innovative vs. non-innovative design within a virtual internship? Specifically, do innovative teams report more frequently that they spend the most time on certain design activities (grouped using Dym’s design framework) versus non-innovative teams? Further, do innovative teams make their final design justifications on the basis of different factors than non-innovative teams do? This research was conducted with sophomore chemical engineering students in the spring 2014 and 2015 semesters. A total of 50 teams of approximately 4-5 students each were studied. Student design processes were evaluated based on innovativeness as well as weekly journal entries where students reported the three activities they spent the most time on. Our results showed no significant differences between innovative and non-innovative teams in terms of their reports of the Dym’s-based activities that they spent the most time on, although our sample size was small. The Management category was associated with the largest effect size (d=0.68), with innovative teams reporting more frequently than non-innovative teams that they spent the most time on design activities that were Management-related. In terms of attributes that contributed to innovative products, teams with higher innovation scores tended to prioritize cost and membrane efficiency (as determined by maximum allowable flux) over patient comfort (measured by blood cell reactivity) and projected market sales. Our results are intended to provide a map for design processes that may ultimately lead to more innovative designs within a virtual internship environment

A Grounded Qualitative Analysis of the Effect of a Focus Group on Design Process in a Virtual Internship

A key component associated with the development of an entrepreneurial mindset is the ability to understand customerneeds and consider this when developing a product. This study sought to understand whether the inclusion of a customerfocus group as part of a virtual internship created any differences in the design processes of sophomore engineeringstudents (114 students). The Nephrotex virtual internship requires that students design a dialysis membrane by optimizinga selection of four components: membrane polymer, polymerization process, processing surfactant, and carbon nanotubepercentage. We found that sophomores who engaged in a focus group during the virtual internship Nephrotex showed(statistically) equal focus on cost versus technical measures of design performance during the focus group. Despite this,design cost was lower in the section that participated in a focus group, with no decrease in product quality. This indicatesthat customer voice may be an important factor in decreasing product cost. We also found that sophomore studentsprioritized their interviewing of customers within the focus group towards end users, such as the patient and nephrologist.Qualitative analysis of sophomore responses demonstrated that they found utility in the focus group (30% of participants)but did not necessarily believe that the customers had useful knowledge of the relevant design attributes (17% ofparticipants). Such realizations may have contributed to the equivalent quality and decreased costs associated with thedesigns of sophomores who participated in a focus group

Normalizing salt content by mixing native human airway mucus samples normalizes sample rheology

Across the globe, millions of people are affected by muco-obstructive pulmonary diseases like cystic fibrosis, asthma, and chronic obstructive pulmonary disease. In MOPDs, the airway mucus becomes hyperconcentrated, increasing viscoelasticity and impairing mucus clearance. Research focused on treatment of MOPDs requires relevant sources of airway mucus both as a control sample type and as a basis for manipulation to study the effects of additional hyperconcentration, inflammatory milieu, and biofilm growth on the biochemical and biophysical properties of mucus. Endotracheal tube mucus has been identified as a prospective source of native airway mucus given its several advantages over sputum and airway cell culture mucus such as ease of access and in vivo production that includes surface airway and submucosal gland secretions. Still, many ETT samples suffer from altered tonicity and composition from either dehydration, salivary dilution, or other contamination. Herein, the biochemical compositions of ETT mucus from healthy human subjects were determined. Samples were characterized in terms of tonicity, pooled, and restored to normal tonicity. Salt-normalized ETT mucus exhibited similar concentration-dependent rheologic properties as originally isotonic mucus. This rheology agreed across spatial scales and with previous reports of the biophysics of ETT mucus. This work affirms previous reports of the importance of salt concentration on mucus rheology and presents methodology to increase yield native airway mucus samples for laboratory use and manipulation

Pathological mucus and impaired mucus clearance in cystic fibrosis patients result from increased concentration, not altered pH

Cystic fibrosis (CF) is a recessive genetic disease that is characterised by airway mucus plugging and reduced mucus clearance. There are currently alternative hypotheses that attempt to describe the abnormally viscous and elastic mucus that is a hallmark of CF airways disease, including: 1) loss of CF transmembrane regulator (CFTR)-dependent airway surface volume (water) secretion, producing mucus hyperconcentration-dependent increased viscosity, and 2) impaired bicarbonate secretion by CFTR, producing acidification of airway surfaces and increased mucus viscosity. A series of experiments was conducted to determine the contributions of mucus concentration versus pH to the rheological properties of airway mucus across length scales from the nanoscopic to macroscopic. For length scales greater than the nanoscopic, i.e. those relevant to mucociliary clearance, the effect of mucus concentration dominated over the effect of airway acidification. Mucus hydration and chemical reduction of disulfide bonds that connect mucin monomers are more promising therapeutic approaches than alkalisation

An improved inhaled mucolytic to treat airway muco-obstructive diseases

Rationale: Airways obstruction with thick, adherent mucus is a pathophysiologic and clinical feature of muco-obstructive respiratory diseases, including chronic obstructive pulmonary disease, asthma, and cystic fibrosis (CF). Mucins, the dominant biopolymer in mucus, organize into complex polymeric networks via the formation of covalent disulfide bonds, which govern the viscoelastic properties of the mucus gel. For decades, inhaled N-acetylcysteine (NAC) has been used as a mucolytic to reduce mucin disulfide bonds with little, if any, therapeutic effects. Improvement of mucolytic therapy requires the identification of NAC deficiencies and the development of compounds that overcome them. Objectives: Elucidate the pharmacological limitations of NAC and test a novel mucin-reducing agent, P3001, in preclinical settings. Methods: The study used biochemical (e.g., Western blotting, mass spectrometry) and biophysical assays (e.g., microrheology/macrorheology, spinnability, mucus velocity measurements) to test compound efficacy and toxicity in in vitro and in vivo models and patient sputa. Measurements and Main Results: Dithiothreitol and P3001 were directly compared with NAC in vitro and both exhibited superior reducing activities. In vivo, P3001 significantly decreased lung mucus burden in bENaC-overexpressing mice, whereas NAC did not (n = 6–24 mice per group). In NAC-treated CF subjects (n = 5), aerosolized NAC was rapidly cleared from the lungs and did not alter sputum biophysical properties. In contrast, P3001 acted faster and at lower concentrations than did NAC, and it was more effective than DNase in CF sputum ex vivo. Conclusions: These results suggest that reducing the viscoelasticity of airway mucus is an achievable therapeutic goal with P3001 class mucolytic agents

Endotracheal tube mucus as a source of airway mucus for rheological study

Muco-obstructive lung diseases (MOLDs), like cystic fibrosis and chronic obstructive pulmonary disease, affect a spectrum of subjects globally. In MOLDs, the airway mucus becomes hyperconcentrated, increasing osmotic and viscoelastic moduli and impairing mucus clearance. MOLD research requires relevant sources of healthy airway mucus for experimental manipulation and analysis. Mucus collected from endotracheal tubes (ETT) may represent such a source with benefits, e.g., in vivo production, over canonical sample types such as sputum or human bronchial epithelial (HBE) mucus. Ionic and biochemical compositions of ETT mucus from healthy human subjects were characterized and a stock of pooled ETT samples generated. Pooled ETT mucus exhibited concentration-dependent rheologic properties that agreed across spatial scales with reported individual ETT samples and HBE mucus. We suggest that the practical benefits compared with other sample types make ETT mucus potentially useful for MOLD research