Pulmonary Artery Remodeling and Advanced Hemodynamics: Magnetic Resonance Imaging Biomarkers of Pulmonary Hypertension

Featured Application: Pulmonary artery remodeling and 3D ﬂow biomarkers can be useful to characterize pulmonary hypertension severity and progression. Abstract: Poorly characterized by non-invasive diagnostic imaging techniques, pulmonary hypertension (PHT) is commonly associated with changes in vascular hemodynamics and remodeling of pulmonary artery architecture. These disease phenotypes represent potential biomarkers of interest in clinical environment. In this retrospective clinical study, 33 patients with pulmonary hypertension and seventeen controls were recruited. Architectural remodeling was characterized using 3D-contrast enhanced angiogram via the measurement of pulmonary artery diameters, bifurcation distances, and angles. Hemodynamics were characterized using 4D-ﬂow magnetic resonance imaging (MRI) via wall shear stress, kinetic energy, vorticity, and directional ﬂow dynamics. Parameters were compared using independent samples student’s t-tests. Correlational analysis was performed using Pearson’s correlation. PHT patients demonstrated dilation in the main and right branch of the pulmonary artery ( p < 0.05). Furthermore, these patients also exhibited increases in bifurcation distances in the left and right pulmonary arteries ( p < 0.05). Wall shear stress, maximum kinetic energy, and energy loss were decreased in the pulmonary artery ( p < 0.001). Correlations were observed between peak velocities and right ventricle ejection fraction (r = 0.527, p < 0.05). These ﬁndings suggest that pulmonary artery remodeling and hemodynamic changes may possess clinical utility as MRI biomarkers for PHT. Author Contributions: Conceptualization, Z.M.H. and J.G.; methodology, J.G.; software, J.G.; validation, Z.M.H. and J.G.; formal analysis, Z.M.H. and J.G.; investigation, Z.M.H. and J.G.; resources, J.G.; data curation, Z.M.H. and J.G.; writing—original draft preparation, Z.M.H. and J.G.; writing—review and editing, J.G.; visualization, Z.M.H. and J.G.; supervision, J.G.; project administration, J.G.; funding


Introduction
Pulmonary hypertension (PHT) is a complex pathophysiological disorder characterized by an elevation in pulmonary arterial pressure (PAP). This diagnosis is given when mean PAP exceeds 25 mmHg at rest using invasive right heart catheterization [1]. In early stages of the disease, elevations in PAP and architectural remodeling typically induce dilation and overload-induced hypertrophy of the right ventricle. Over time, this results in impairment of cardiac function, eventual right heart failure, and secondary pathophysiology such as right atrial dilation [2]. Although two-dimensional (2D) Doppler echocardiography is commonly used to provide evidence of the disease hallmarks in the right ventricle, and right ventricular catherization is needed for a conclusive diagnosis. Thus, other non-invasive diagnostic imaging biomarkers would be beneficial in clinical settings [3]. Magnetic resonance imaging (MRI) and angiography (MRA) could address this unmet need. Relative to 2D Doppler echocardiography, MRI possesses two advantages.
A total of 51 subjects were recruited retrospectively, including 17 PHT patients and 33 controls. All subjects were registered within the Cardiovascular Imaging Registry of Calgary (CIROC). The study was approved by the University of Calgary Research Ethics Board and all subjects provided written informed consent. All research activities were performed in accordance with the Declaration of Helsinki. The study was coordinated by commercial software (cardioDI TM , Cohesic Inc., Calgary, AB, Canada) for the routine capture of patient informed consent, for health questionnaires, and for standardized collection of MRI-related variables. PHT was clinically defined as a mean PA pressure ≥25 mmHg previously assessed by clinical invasive catheterization or echocardiography. Patients were recruited at the time of visit, with exclusions occurring for patients with prior surgical interventions in the pulmonary valve and/or artery; complex congenital heart disease; and/or contraindications to MRI. Control subjects (≥18 years of age) underwent similar workflow and confirmed no prior history of cardiovascular disease with a certified nurse. No catherization was performed in healthy controls. Prior to scanning, demographic measurements including age, sex, height, weight, and heart rate were obtained. Volume and mass measurements were normalized to body surface area using the Mosteller formula.

Cardiac Magnetic Resonance Imaging Protocol
Cardiac imaging examination was performed using 3T MRI scanners (Skyra and Prima, Siemens, Erlangen, Germany). Indication-based protocolling ensured consistent imaging procedures for all subjects, and cardiac imaging was performed in accordance with published recommendations [9]. Standard routine retrospective electrocardiographic gating, and timeresolved balanced steady-state free precession (SSFP) cine imaging in four-chamber, threechamber, two-chamber, and short-axis views of LV at end-expiration, was performed. Contrast usage of gadolinium contrast volume of 0.2 mmol/kg (Gadovist ® , Bayer Inc., Mississauga, ON, Canada) was administrated to acquire a contrast-enhanced 3D magnetic resonance angiogram (CE MRA) of the cardiovascular structure. Time-resolved three-dimensional phase-contrast MRI with three-directional velocity encoding and retrospective ECG-gating (4D-flow, Siemens WIP 785A) was performed for 5-10 min, following contrast administration to measure in-vivo blood flow velocities within the whole heart. We have previously reported this whole-heart protocol [10][11][12]. Briefly, 4D-flow data was acquired during free breathing using navigator gating of diaphragmatic motion; sequence parameters were as follows: flip angle = 15 degrees, spatial resolution = 2.0−3.5 × 2.0−3.5 × 2.5−3.5 mm; temporal resolution = 39-48 ms; and velocity sensitivity = 150−250 cm/s. Total acquisition time varies between 5-10 min, depending on heart rate and respiratory navigator efficiency. The number of phases was adjusted to 25.

Standard Cardiac Imaging Analysis
Standard cardiac images were analyzed by a blinded reader to the study the same day of the acquisition using dedicated software cvi 42 version 5.11.5 (Circle Cardiovascular Imaging Inc, Calgary, AB, Canada) to determine left and right end-diastolic volume (LVEDV; RVEDV), LV and RV end-systolic volume (LVSEV; RVESV), and LV and RV ejection fraction (LVEF; RVEF). Furthermore, the contrast-enhanced angiogram (CE MRA) was used to measure pulmonary artery diameters (LPA, MPA, and RPA), bifurcation distances, and bifurcation angles ( Figure 1D). navigator gating of diaphragmatic motion; sequence parameters were as follows: flip angle = 15 degrees, spatial resolution = 2.0−3.5 × 2.0−3.5 × 2.5−3.5 mm; temporal resolution = 39-48 ms; and velocity sensitivity = 150−250 cm/s. Total acquisition time varies between 5-10 min, depending on heart rate and respiratory navigator efficiency. The number of phases was adjusted to 25.

Standard Cardiac Imaging Analysis
Standard cardiac images were analyzed by a blinded reader to the study the same day of the acquisition using dedicated software cvi 42 version 5.11.5 (Circle Cardiovascular Imaging Inc, Calgary, AB, Canada) to determine left and right end-diastolic volume (LVEDV; RVEDV), LV and RV end-systolic volume (LVSEV; RVESV), and LV and RV ejection fraction (LVEF; RVEF). Furthermore, the contrast-enhanced angiogram (CE MRA) was used to measure pulmonary artery diameters (LPA, MPA, and RPA), bifurcation distances, and bifurcation angles ( Figure 1D).

4D-Flow Data Analysis
All 4D-flow MRI data was pre-processed using in-house program developed in MAT-LAB 2020b (Mathworks, Natick, MA, USA), and the following tasks were performed: corrections for Maxwell terms, eddy currents, and aliasing, Figure 1A. After pre-processing, a 3D phase-contrast (PC) angiogram (PC MRA) was generated, Figure 1B. This angiogram was used to segment selected anatomical regions of the pulmonary artery, Figure 1B red inset, using in-house MATLAB-based tool "4D-Flow Analysis Tool" [13,14]. Analysis planes were created at the right ventricular outflow tract (RVOT), main pulmonary artery (MPA), the left pulmonary artery (LPA), and right pulmonary artery (RPA) using a specialized visualization software (Ensight 10.2, CEI Inc, Research Triangle Park, North Carolina, USA) to measure peak velocity, net flow, retrograde flow, regurgitation fraction, and forward flow, Figure 1C. Furthermore, the PC MRA was used to measure pulmonary artery diameters (LPA, MPA and RPA), bifurcation distances, and bifurcation angles ( Figure 1D) in cvi 42 when standard CE MRA was not available in controls (n = 2). Volume sub-regions were created for local advanced 4D-flow analysis. These volume subregions were used to obtain the maximum velocities from the LPA, PA, and RPA using the approach proposed by Rose et al. [15].
Regional WSS was calculated at peak systole as previously described [16,17]. Energy loss (EL) was calculated as proposed by Barker et al. [18], which provides the rate of EL in a volume of interest (i.e., power in Watts) at a given time of the cardiac cycle, and peak systole was used in this study. Kinetic energy (KE) was calculated as reported by Geeraert et al. [13] in each sub-region at peak systole. Vortex size analysis at peak systole was performed as introduced by Garcia et al. [10] using the Lambda2 (λ 2 ) method, which is an accepted method to identify 3D vortices. Valid vortex networks were defined as those >75 mm 3 (considering a voxel resolution of 2.5 × 2.5 × 3.0 mm) with a shared connectivity neighborhood ≥4 voxels. This rule reduces the impact of possible incorrect derived vortices due to noise and/or partial derivative errors, which are directly impacted by 4D-flow spatial resolution. Examples for these advanced 4D-flow parameters are presented in Figure 1E.

Statistical Analysis
Statistical analysis was performed using SPSS 25 (SPSS, Chicago, IL, USA). Normality was assessed using normal plots and the Shapiro-Wilk test. Between-group comparisons for demographic, baseline cardiac MRI, hemodynamics, and anatomical measurements were performed using an independent samples t-test incorporating Levene's test for equality of variances. To assess the relationship between variables, a series of linear regression tests was performed using Pearson R-Values. If a significant relationship was found, a univariate linear model was generated using variables demonstrating statistical significance during the linear regression tests. Pearson correlation coefficients were also generated in relation to LVEF and RVEF. The significance threshold was <0.05.

Univariate Analysis
Correlational analyses were reported in Table 4

Discussion
In this study, we demonstrate that PHT is associated with both hemodynamic and anatomic changes throughout the PA. Patients with PHT demonstrated multiple changes across a variety of hemodynamic parameters, including EL, WSS, vorticity, flow direction and flow velocity. In addition, pulmonary artery dilation was observed alongside changes in bifurcation geometry. Sex matching for women identified differences for several anatomical and advanced hemodynamic biomarkers. Taken together, these findings provide additional evidence that hemodynamic and anatomic biomarkers possess clinical relevance for diagnosis of the disease.
Although hemodynamic changes are emerging biomarkers of the disease, they remain poorly understood. Previous research suggests patients with PHT demonstrate lower wall shear stress [5,7]. Although these studies were conducted using an adult PHT population, the findings possess relevance to the broader definition of PHT. For vorticity, a recent study conducted by Kroeger et al. found that vortices are observed in healthy controls and patients with PHT [19]. Similarly, Schafer et al. identified reduced vorticity but increased helicity in patients with PHT [6]. For EL, Han et al. demonstrated greater total energy losses over the cardiac cycle throughout the entirety of the PA [20] in PHT patients. PHT patients may also demonstrate greater retrograde flow and lower peak velocity [21,22]. In contrast, anatomical changes are more well-studied using CT, identifying dilation of the main pulmonary artery in various stages of the disease [23,24]. These findings have been replicated using MRI and appear to be prevalent in the later stages [25].
Our findings generally corroborate results of previous studies and provide insight into novel biomarkers. We observed lower WSS, maximum energy attained, and regurgitation fractions in the LPA and RPA in patients with PHT. Maximum EL at peak systole remained lower in patients than in controls in all PA sections. Mean EL tended to be also lower, except for the LPA. EL reported in our study was quantified at peak systole, which provides further support in addition of previous findings reported over the cardiac cycle. Contrary to other studies, we found that vorticity tended to increase in PHT patients. For anatomical changes, we observed dilation in the MPA and part of the RPA. Of note, we are the first study to our knowledge to identify greater bifurcation distances associated with PHT. We speculate this could be an additional form of anatomical remodeling associated with disease progression. However, the exact significance is unknown. Further studies investigating changes in bifurcation geometry at various disease stages are needed to better understand their significance. The difference between the RPA and LPA characterization may be influenced by the patient's anatomy and the hemodynamic resistance from the lungs. However, the latter requires additional investigation. Furthermore, sex-matching analysis confirmed the impact of advanced hemodynamic parameters in PHT patients and highlighted the importance of subject matching in future studies.
This study has a few limitations. First, our study was limited by the spatiotemporal resolution constraints of MRI. For example, while we could visualize larger vortex networks, smaller vortices occurring at faster speeds exceeding the available resolution could be missed. Second, as a single centre study, the addition of multiple centres would improve the generalization of results. Furthermore, due to its retrospective nature, the sample size and power is limited. Third, our control patients were not age and sex-normalized to the patient cohort, which suggests normal cardiac aging and sex differences could have impacted results. Fourth, PHT is a progressive multi-factorial condition with diverse underlying etiology. Clinical protocols did not mandate further characterization of the disease into smaller subgroups such as pulmonary arterial hypertension, or patients at varying stages of the disease, limiting the characterization of disease progression. The relatively small cohort size did not allow one to explore the effect of medical therapy in 4D-flow derived parameters. The latter remains an important aspect to assess in future studies. Finally, we did not explore the right ventriculo-pulmonary arterial coupling in the present cohort. This coupling analysis may produce a better understanding of the bidirectional effects between the RV and the pulmonary artery tree.

Conclusions
Adult PHT patients demonstrate hemodynamic and anatomic differences throughout the PA. In addition to providing further support for established disease phenotypes such as reduced WSS and PA dilatation, these findings suggest that novel biomarkers such as energy changes and changes in bifurcation geometry may possess clinical relevance for PHT patients. Future studies are needed to investigate these phenomena at varying stages of the disease and determine how they may be affected by underlying etiology. Informed Consent Statement: Written informed consent was obtained from all subjects involved in the study.

Data Availability Statement:
The anonymized data presented in this study are available on request from the corresponding author. The data are not publicity available due to privacy and ethical restrictions.

Conflicts of Interest:
The authors declare no conflict of interest.