Search

Phase contrast x-ray imaging (PCXI) provides high-contrast images of weakly-attenuating structures like the lungs. PCXI, when paired with 4D X-ray Velocimetry (XV), can measure regional lung function and non-invasively assess the efficacy of emerging therapeutics. Bacteriophage therapy is an emerging antimicrobial treatment option for lung diseases such as cystic fibrosis (CF), particularly with increasing rates of multi-drug-resistant infections. Current efficacy assessment in animal models is highly invasive, typically requiring histological assessment. We aim to use XV techniques as non-invasive alternatives to demonstrate efficacy of bacteriophage therapy for treating Pseudomonas aeruginosa CF lung infections, measuring functional changes post-treatment. Time-resolved in vivo PCXI-CT scans of control, Pseudomonas-infected, and phage-treated mouse lungs were taken at the Australian Synchrotron Imaging and Medical Beamline. Using XV we measured local lung expansion and ventilation throughout the breath cycle, analysing the skew of the lung expansion distribution. CT images allowed visualisation of the projected air volume in the lungs, assessing structural lung damage. XV analysis demonstrated changes in lung expansion between infection and control groups, however there were no statistically significant differences between treated and placebo groups. In some cases where structural changes were not evident in the CT scans, XV successfully detected changes in lung function., Competing Interests: Declaration. Competing interests: Martin Donnelley and Stephen Dubsky own shares in 4DMedical. Stephanie A. Harker, Melissa Preissner, Rachel Yoon Chang, David Trevascus, Chengxi Liu, Yuncheng Wang, Michael Yee Chow, Patricia Cmielewski, Nicole Reyne, Ying Ying How, James A. Pollock, Mitzi Klein, Christopher A. Wright, Hak-Kim Chan, and Kaye S. Morgan have no competing interests., (© 2024. The Author(s).)

To effectively diagnose, monitor and treat respiratory disease clinicians should be able to accurately assess the spatial distribution of airflow across the fine structure of lung. This capability would enable any decline or improvement in health to be located and measured, allowing improved treatment options to be designed. Current lung function assessment methods have many limitations, including the inability to accurately localise the origin of global changes within the lung. However, X-ray velocimetry (XV) has recently been demonstrated to be a sophisticated and non-invasive lung function measurement tool that is able to display the full dynamics of airflow throughout the lung over the natural breathing cycle. In this study we present two developments in XV analysis. Firstly, we show the ability of laboratory-based XV to detect the patchy nature of cystic fibrosis (CF)-like disease in β-ENaC mice. Secondly, we present a technique for numerical quantification of CF-like disease in mice that can delineate between two major modes of disease symptoms. We propose this analytical model as a simple, easy-to-interpret approach, and one capable of being readily applied to large quantities of data generated in XV imaging. Together these advances show the power of XV for assessing local airflow changes. We propose that XV should be considered as a novel lung function measurement tool for lung therapeutics development in small animal models, for CF and for other muco-obstructive diseases.

Most measures of lung health independently characterise either global lung function or regional lung structure. The ability to measure airflow and lung function regionally would provide a more specific and physiologically focused means by which to assess and track lung disease in both pre-clinical and clinical settings. One approach for achieving regional lung function measurement is via phase contrast X-ray imaging (PCXI), which has been shown to provide highly sensitive, high-resolution images of the lungs and airways in small animals. The detailed images provided by PCXI allow the application of four-dimensional X-ray velocimetry (4DxV) to track lung tissue motion and provide quantitative information on regional lung function. However, until recently synchrotron facilities were required to produce the highly coherent, high-flux X-rays that are required to achieve lung PCXI at a high enough frame rate to capture lung motion. This paper presents the first translation of 4DxV technology from a synchrotron facility into a laboratory setting by using a liquid-metal jet microfocus X-ray source. This source can provide the coherence required for PCXI and enough X-ray flux to image the dynamics of lung tissue motion during the respiratory cycle, which enables production of images compatible with 4DxV analysis. We demonstrate the measurements that can be captured in vivo in live mice using this technique, including regional airflow and tissue expansion. These measurements can inform physiological and biomedical research studies in small animals and assist in the development of new respiratory treatments.

The beating heart is known to produce pressure and airflow oscillations in the lungs of mammals. This phenomenon is often disregarded as detailed measurement of its effects in the lung have hitherto not been possible. Previous studies have attempted to measure the effect of these oscillations on gas mixing. However, the results have proven inconclusive, due to the lack of a direct measurement tool capable of flow measurement throughout the entire bronchial tree. Here we present the first detailed measurement of cardiogenic oscillations, using synchrotron-based dynamic lung imaging of live mechanically ventilated mice. The results demonstrate large flow oscillations and pendelluft in the airways due to the mechanical action of the beating heart. Using a virtual tracer modelling analysis we show that cardiogenic oscillations produced up to 4 times increased gas mixing, but only in the absence of tidal ventilation. The results highlight the importance of considering this often-disregarded phenomenon when investigating lung function, particularly in situations where tidal ventilation is reduced or absent.

Computed tomography (CT) and spirometry are the mainstays of clinical pulmonary assessment. Spirometry is effort dependent and only provides a single global measure that is insensitive for regional disease, and as such, poor for capturing the early onset of lung disease, especially patchy disease such as cystic fibrosis lung disease. CT sensitively measures change in structure associated with advanced lung disease. However, obstructions in the peripheral airways and early onset of lung stiffening are often difficult to detect. Furthermore, CT imaging poses a radiation risk, particularly for young children, and dose reduction tends to result in reduced resolution. Here, we apply a series of lung tissue motion analyses, to achieve regional pulmonary function assessment in β-ENaC-overexpressing mice, a well-established model of lung disease. The expiratory time constants of regional airflows in the segmented airway tree were quantified as a measure of regional lung function. Our results showed marked heterogeneous lung function in β-ENaC-Tg mice compared to wild-type littermate controls; identified locations of airway obstruction, and quantified regions of bimodal airway resistance demonstrating lung compensation. These results demonstrate the applicability of regional lung function derived from lung motion as an effective alternative respiratory diagnostic tool.