Chronic Thromboembolic Pulmonary Hypertension Model Research Articles

Suppression of Aseptic Inflammation Reduces the Severity of Remodeling of the Pulmonary Artery Branches and Improves Progressing of Experimental Chronic Thromboembolic Pulmonary Hypertension

Chronic thromboembolic pulmonary hypertension (CTEPH) is a complication of pulmonary embolism, characterized by increased pressure in the pulmonary artery and impaired lysis of thromboemboli. Previously, the presence of aseptic inflammation in CTEPH was identified in the wall of the pulmonary artery branches and perivascularly. However, the role of this inflammation in the CTEPH formation is unknown. The aim of the work was to study the effect of aseptic inflammation on the CTEPH formation and progression. The experiments were performed on 54 male rats. The CTEPH model was reproduced by repeated intravenous administration of partially biodegradable microspheres (MS). Immediately after the last administration of MS, all animals were divided into groups: control CTEPH (c.CTEPH) – saline solution was administered intramuscularly (i/m) for 6 weeks; low dose of prednisolone (LD) – prednisolone was administered i/m at a dose of 1.5 mg/kg; high dose (HD) – prednisolone was administered i/m at a dose of 6 mg/kg; healthy animals. After 6 weeks, the following was performed: treadmill test, TTE, cardiac catheterization with manometry, and histological examination of the lungs. In a separate series of experiments, the severity of inflammatory infiltration of the vascular wall and perivascular zone was assessed by immunohistochemical studies (IHC). In the LD group, there was the decreasing of hypertrophy index (HI) and the percentage of collagen fibers in the vascular wall compared to c.CTEPH. There was a significantly greater reduction in HI compared to HD. In the HD group, there was positive effect on the percentage of collagen fibers in the vascular wall, this parameter did not significantly differ from healthy animals. According to IHC data, prednisolone in low dose effectively suppressed inflammatory infiltration of the vascular wall and perivascular space. The results of the study revealed the ability of prednisolone, by suppressing aseptic inflammation, to reduce the severity of remodeling of the pulmonary artery branches.

Subject-specific one-dimensional fluid dynamics model of chronic thromboembolic pulmonary hypertension

Chronic thromboembolic pulmonary hypertension (CTEPH) develops due to the accumulation of blood clots in the lung vasculature that obstructs flow and increases pressure. The mechanobiological factors that drive progression of CTEPH are not understood, in part because mechanical and hemodynamic changes in the small pulmonary arteries due to CTEPH are not easily measurable. Using previously published hemodynamic measurements and imaging from a large animal model of CTEPH, we applied a subject-specific one-dimensional (1D) computational fluid dynamic (CFD) approach to investigate the impact of CTEPH on pulmonary artery stiffening, time-averaged wall shear stress (TAWSS), and oscillatory shear index (OSI) in extralobar (main, right, and left) pulmonary arteries and intralobar (distal to the extralobar) arteries. Our results demonstrate that CTEPH increases pulmonary artery wall stiffness and decreases TAWSS in extralobar and intralobar arteries. Moreover, CTEPH increases the percentage of the intralobar arterial network with both low TAWSS and high OSI, quantified by the novel parameter φ\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\varphi$$\\end{document}, which is related to thrombogenicity. Our analysis reveals a strong positive correlation between increases in mean pulmonary artery pressure (mPAP) and φ\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\varphi$$\\end{document} from baseline to CTEPH in individual subjects, which supports the suggestion that increased φ\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\varphi$$\\end{document} drives disease severity. This subject-specific experimental–computational framework shows potential as a predictor of the impact of CTEPH on pulmonary arterial hemodynamics and pulmonary vascular mechanics. By leveraging advanced modeling techniques and calibrated model parameters, we predict spatial distributions of flow and pressure, from which we can compute potential physiomarkers of disease progression. Ultimately, this approach can lead to more spatially targeted interventions that address the needs of individual CTEPH patients.

The Use of Microencapsulated Autologous Thrombi for Modelling Chronic Thromboembolic Pulmonary Hypertension in Rats.

Here we developed a model of chronic thromboembolic pulmonary hypertension (CTEPH) using repeated intravenous administration of microencapsulated thrombi with a controlled rate of biodegradation. Autologous thrombi encapsulated in alginate microspheres with a diameter of 190±48 μm were intravenously injected to rats 8 times every 4 days. In the comparison group, nonmodified thrombi were injected. After 6 weeks, a significant increase in systolic pressure in the right ventricle, a decrease in exercise tolerance, and an increase in the index of vascular wall hypertrophy were revealed in the group receiving injections of microencapsulated thrombi in comparison with the group receiving nonmodified thrombi and healthy animals. Thus, the developed representative CTEPH model can be used to test promising pharmacological substances.

Overexpressed microRNA (miR)-382-3p promoted vascular remodeling via suppressing autophagy-related protein 7 (ATG7) in chronic thromboembolic pulmonary hypertension.

This research has investigated the role of miR-382-3p in chronic thromboembolic pulmonary hypertension (CTEPH). Human pulmonary artery smooth muscle cells (hPASMCs) were treated with PDGF-BB to induce proliferation, and then transfected with miR-382-3p mimic, miR-382-3p inhibitor, ATG7 overexpression plasmid, and siATG7. MiR-382-3p, ATG7, VEGF, PCNA, p62, and LC3-Ⅱ/LC3-I levels were detected by qRT-PCR and Western blotting. Cell viability and migration were tested through CCK-8 and wound healing assays, respectively. Target genes of miR-382-3p were predicted by Targetscan and starBase, and pathway analysis was implemented through WebGestalt. The binding relationship between miR-382-3p and ATG7 was analyzed by the dual-luciferase reporter and RIP assays. A CTEPH model was constructed in rats with the treatment of miR-382-3p antagomir or agomir, and mean pulmonary artery pressure (mPAP) was measured. Lung tissue was observed through the HE staining assay. MiR-382-3p level in hPASMCs was obviously upregulated with the increasing dose of PDGF-BB. MiR-382-3p mimic promoted yet miR-382-3p inhibitor suppressed hPASMC proliferation. MiR-382-3p directly targeted ATG7. ATG7 overexpression repressed hPASMC proliferation and migration, whereas siATG7 exerted the opposite effects. ATG7 overexpression partly neutralized the effects of miR-382-3p mimic on proliferation, migration, and autophagy-related proteins (ATG7, p62, and LC3-Ⅱ/LC3-I) in hPASMCs, whereas siATG7 partly offset the impacts of miR-382-3p inhibitor. MiR-382-3p antagomir reversed CTEPH-induced mPAP elevation, miR-382-3p upregulation, thickening of the pulmonary artery wall, and increased expressions of VEGF, PCNA, and autophagy-related proteins in rats, while miR-382-3p agomir potentiated these effects induced by CTEPH. Overexpressed miR-382-3p promotes vascular remodeling via ATG7 inhibition in CTEPH.

Study of the JAK inhibitors antifibrotic activity for the prevention and treatment of chronic thromboembolic pulmonary hypertension

Introduction. Chronic thromboembolic pulmonary hypertension (CTEPH) is the most common complication of pulmonary thromboembolism (PE). Fibrous remodeling of the pulmonary circulation vessels against the background of CTEPH leads to an irreversible increase of the vessel wall stiffness and the ineffectiveness of CTEPH treatment. The involvement of Janus kinase (JAK) in the regulation of vascular wall and lung tissue inflammation and fibrosis allows for the possible effectiveness of JAK 1,2 inhibitors (iJAK) in the course of CTEPH. Purpose – to study the antifibrotic effect of iJAK for the prevention and treatment of CTEPH. Materials and methods. The study was conducted on male Wistar rats. Modeling of CTEPH was performed by sequential embolization of the vascular bed with partially biodegradable sodium alginate microspheres. 2 weeks after the last administration of the microspheres, low, medium and high doses of iJAK were initiated. To assess the effectiveness of the substance, the following tests were used: treadmill test, echocardiography, cardiac catheterization with right ventricular (RV) manometry, histological examination of the lungs. Results. Animals undergone vascular embolization demonstrated decreased exercise tolerance at all observation points compared to healthy animals. The placebo group, in contrast with the group getting treatment and iJAK, was found to have an increased mean RV pressure compared to healthy animals. There was an increase in mean RV pressure in the placebo group (15.5±7.7 mmHg) and in the low dose and iJAK group (13.4±6.4 mmHg) compared with healthy animals (9.4±2.2 mmHg). Vascular hypertrophy of the pulmonary artery branches was lower in group getting average dosages and iJAK compared with the placebo group (54.9±19.0 and 68.9±23.1 %, respectively). Thus, the suppression by iJAK of aseptic inflammation and following fibrosis leads to the decreasing of severity of pulmonary circulation remodeling in the experimental model of CTEPH. This approach can be used in the comprehensive bypass and prevention of CTEPH.

Metformin Improves Pulmonary Artery Remodeling in Chronic Thromboembolic Pulmonary Hypertension Rats

Abstract Background To investigate the effect of metformin on pulmonary artery remodeling and indexes of endothelial–mesenchymal transition in rats with chronic thromboembolic pulmonary hypertension (CTEPH). Methods Sixty Sprague Dawley rats were randomly divided into normal control, metformin control, CTEPH, metformin low-dose (50 mg/kg d) CTEPH, and metformin high-dose group (100 mg/kg d) CTEPH groups. The autologous blood clots were repeatedly injected through the jugular vein to establish the CTEPH model. After 6 weeks treatment with or without metformin, the mean pulmonary artery pressure (mPAP), pulmonary artery wall area to total tube area ratio (WA%), and protein levels of vimentin, α-smooth muscle actin (α-SMA), CD31, E-cadherin, β-catenin, and p-adenosine 5′-monophosphate activated protein kinase (p-AMPK) in the pulmonary artery were measured. Results The mPAP and WA% were significantly higher in CTEPH compared with normal control rats (P < 0.05). Metformin treatment dose dependently decreased mPAP and WA% in CTEPH rats (P < 0.01). Compared with normal controls, protein levels of vimentin, α-SMA, and β-catenin were significantly increased while CD31 and E-cadherin levels were decreased in CTEPH rats (all P < 0.05). There was no significant difference in p-AMPK levels between normal control and CTEPH groups. Metformin treatment increased p-AMPK (P < 0.05) but had no significant effect on vimentin, α-SMA, CD31, E-cadherin, or β-catenin levels in control rats. Metformin treatment dose dependently decreased vimentin, α-SMA, and β-catenin levels, and increased CD31, E-cadherin, and p-AMPK levels in CTEPH rats (P < 0.05). Conclusions Metformin reduces mPAP and improves pulmonary vascular remodeling possibly through attenuating endothelial–mesenchymal transition of the pulmonary artery in CTEPH rats.

Left Ventricular Diastolic Dysfunction in Chronic Thromboembolic Pulmonary Hypertension

<h3>Purpose</h3> Impairment of left ventricular (LV) relaxation is associated with increased mortality after surgical treatment of pulmonary hypertension (PH). We sought to describe the pathophysiology of LV diastolic dysfunction in a porcine model of chronic thromboembolic pulmonary hypertension (CTEPH). The reversibility of LV diastolic dysfunction was investigated in patients with CTEPH after pulmonary endarterectomy (PEA). <h3>Methods</h3> CTEPH was induced in 2-month-old Large White piglets (PH group, n=6) by ligation of the left pulmonary artery (PA) followed by weekly embolization of right lower lobe for 5 weeks using a strong tissue glue (N-acetyl cyanoacrylate). These animals were compared to sham-operated animals (controls, n=6). LV diastolic function was assessed using echocardiography and conductance catheter measurements. LV fibrosis was investigated at 6 weeks using red Sirius staining of myocardial tissues. Echocardiographic measurements for LV diastolic function were retrospectively analyzed in 102 patients, before and after PEA. <h3>Results</h3> Mean PA pressure was higher at 6 weeks in PH animals compared to controls (28.5 [28.0; 34.2] vs. 14.0 [12.5; 14.0] mmHg, p<0.01). Increased end-diastolic LV pressure was observed in PH group (21.9 [18.1; 22.7] vs. 12.2 [11.7; 13.8] mmHg, p=0.013), along with a marked decrease in the curve-fitting constant (c) and the maximum rate of LV filling (dV/dt_max), respectively by 49% (p=0.03) and 74% (p=0.014). Stiffness constant ß was strongly correlated with Doppler imaging index E/A (r=-0.94, p=0.015). Mean LV fibrosis score was significantly higher in PH group at 6 weeks (5.11±0.89% vs. 3.29±1.14%, p<0.01). Pre-operative impairment of LV filling pattern was remarkable in patients with CTEPH (E/A = 0.81±0.32; E/E'=6.42±2.84), and significantly improved at 7 days post PEA (+30%, p<0.001). At 6 months, E/A ratio was significantly higher (0.91 [0.75; 1.20] vs. 0.76 [0.66; 0.94], p=0.05) but remained abnormal, despite significant decrease in mean pulmonary vascular resistance (7.3 ±3.1WU vs. 3.8±1.5WU, p<0.001). <h3>Conclusion</h3> Impaired myocardial stiffness was associated with LV fibrosis in our piglet model of CTEPH. Mild LV diastolic dysfunction was observed at 6 months in patient with CTEPH despite significant decrease in RV pressure overload after PEA. Myocardial fibrosis may be responsible for persistent abnormal LV relaxation.

Modulation of soluble guanylate cyclase ameliorates pulmonary hypertension in a rat model of chronic thromboembolic pulmonary hypertension by stimulating angiogenesis.

Acute pulmonary embolism (PE) does not always resolve after treatment and can progress to chronic thromboembolic disease (CTED) or the more severe chronic thromboembolic pulmonary hypertension (CTEPH). The mechanisms surrounding the likelihood of PE resolution or progress to CTED/CTEPH remain largely unknown. We have developed a rat model of CTEPH that closely resembles the human disease in terms of hemodynamics and cardiac manifestations. Embolization of rats with polystyrene microspheres followed by suppression of angiogenesis with the inhibitor of vascular endothelial growth factor receptor 2 (VEGF‐R2) SU5416 results in transient, acute pulmonary hypertension that progresses into chronic PE with PH with sustained right ventricular systolic pressures exceeding 70 mmHg (chronic pulmonary embolism [CPE] model). This model is similar to the widely utilized hypoxia/SU5416 model with the exception that the “first hit” is PE. Rats with CPE have impaired right heart function characterized by reduced VO2Max, reduced cardiac output, and increased Fulton index. None of these metrics are adversely affected by PE alone. Contrast‐mediated CT imaging of lungs from rats with PE minus SU5416 show large increases in pulmonary vascular volume, presumably due to an angiogenic response to acute PE/PH. Co‐treatment with SU5416 suppresses angiogenesis and produces the CTEPH‐like phenotype. We report here that treatment of CPE rats with agonists for soluble guanylate cyclase, a source of cGMP which is in turn a signal for angiogenesis, markedly increases angiogenesis in lungs, and ameliorates the cardiac deficiencies in the CPE model. These results have implications for future development of therapies for human CTEPH.

The role of β-catenin in pulmonary artery endothelial-mesenchymal transformation in rats with chronic thromboembolic pulmonary hypertension.

β-catenin and endothelial mesenchymal transformation play an important role in the formation of pulmonary hypertension. To explore the role of β-catenin in chronic thromboembolic pulmonary hypertension (CTEPH), we first established a rat model of CTEPH by repeated autologous thromboembolization and then treated these rats with a β-catenin specific inhibitor, XAV939, for two or four weeks. We further examined the expression of β-catenin, α-SMA and CD31, mean pulmonary artery pressure (mPAP), and histopathology in the pulmonary artery, and analyzed their correlation. In the thrombus group without treatment of the inhibitor, the expression of β-catenin and α-SMA in pulmonary artery was increased with time; mPAP, the thickness of pulmonary artery wall, and the area/total area of pulmonary artery (WA/TA) were also increased; however, the expression of CD31 was decreased. Interestingly, these symptoms could be improved by treatment with XAV939. In this study, in CTEPH rat model, the expression of β-catenin signal affects pulmonary vascular remodeling and pulmonary artery pressure, and positively correlated with pulmonary arterial endothelial mesenchymal transformation (EMT), indicating that β-catenin signal may play an important role in the occurrence and development of CTEPH. The inhibition of β-catenin signal and the improvement of pulmonary arterial EMT may provide therapeutic ideas for CTEPH.

Description, Staging and Quantification of Pulmonary Artery Angiophagy in a Large Animal Model of Chronic Thromboembolic Pulmonary Hypertension.

Angiophagy has been described as a non-fibrinolytic mechanism of pulmonary artery (PA) patency restoration after distal (<50 µm in diameter) pulmonary embolism in mice. We hypothesized that angiophagy could achieve muscularized PA patency restoration after pulmonary embolism in piglets and humans. Angiophagy was defined by pathological assessment as the moving of an embolic specimen from the lumen to the interstitium according to three stages in a pig model of chronic thromboembolic pulmonary hypertension (CTEPH) 6 to 10 weeks after embolization with enbucrilate: the embolic specimen is (I) covered by endothelial cells, (II) covered by endothelial cells and smooth muscle cells, and (III) located in the adventitia. In animals, we observed the three stages of the pulmonary angiophagy of enbucrilate emboli in <300 µm PA. Stages II and III were observed in 300 to 1000 μm PA, and only Stage I was observed in larger-diameter PA (>1000 μm). In lung samples from patients with histories of pulmonary embolisms, we observed PA angiophagy stigma for embolic specimens derived from blood clots and from bone marrow emboli. This study provides an original pathological description and staging of PA angiophagy in a large animal model of CTEPH and in humans after pulmonary embolism.

The role of inflammation in a rat model of chronic thromboembolic pulmonary hypertension induced by carrageenan.

BackgroundChronic thromboembolic pulmonary hypertension (CTEPH) is a life-threatening condition arising from the thrombus and obstructive remodeling of the pulmonary arteries, which causes a significant morbidity and mortality. Although the modern treatment in CTEPH has been significant advanced both in surgical and medical treatment, none can claim to cure the disease, largely because of our limited understanding of the underlying pathogenesis of the disease and lack of a reliable CTEPH animal model to study for. Recently, inflammation has been accepted as a common pathway through which various risk factors trigger venous thrombo-embolism (VTE) formation, we describe a novel mouse model of CTEPH which reproduces a frequent trigger and resembles the time course, histological features, and clinical presentation of CTEPH in humans, to open a new horizons of inflammation in CTEPH.MethodsBy administering a pulmonary embolism (PE) protocol (comprising 3 sequential left jugular vein injections of autologous blood clots) to 8-week-old male Sprague Dawley (SD) rats using tranexamic acid (200 mg/kg.d) to inhibit fibrinolysis and injecting additional carrageenan (20 mg/kg, once a week) to create perivascular inflammation, we successfully generated a CTEPH animal model. By monitoring the mean pulmonary artery pressure (mPAP) and the histopathological change to evaluate the CTEPH model. By detecting the RT-PCR, western blot, TUNEL, and immunohistochemistry in the sub-groups to find the potential mechanism of inflammation may work in the pulmonary vascular remolding.ResultsIn this study, rats with CTEPH exhibited pronounced pulmonary vascular remolding with higher vessel wall area/total area (WA/TA) ratio in comparison to the control rats (85.41%±7.37% vs. 76.41%±5.97%, P<0.05), the mPAP (25.51±1.13 vs. 15.92±1.13 mmHg, P<0.05). Significant differences in mean pulmonary artery pressure (mPAP) values were observed between rats injected solely with clots and those injected with both clots and carrageenan (25.51±1.13 vs. 29.82±1.26 mmHg, P<0.05, respectively). Furthermore, following the third embolization, thrombi and intimal hyperplasia occurred in the pulmonary artery. In addition, repeated embolization elevated mRNA and protein levels of tumor necrosis factor-α (TNF-α), NF-κB/p65, and B-cell lymphoma-2 (BCL-2), but decreased BAX expression in a time-dependent manner.ConclusionsTake advantage of the inflammation to trigger VTE formation, we successfully generated a CTEPH animal model. Inflammation may play a crucial role in the pathogenesis and progression of CTEPH by inhibiting endothelial cell apoptosis. Understanding the role of inflammation in CTEPH may not only help to determine the optimal treatment options but also may aid in the development of future preventative strategies, since current anticoagulation treatment regimens are not designed to inhibit inflammation.

The Protective Effect of The Interleukin 1 Receptor Antagonist on Chronic Thromboembolic Pulmonary Hypertension Model

Chronic thromboembolic pulmonary hypertension (CTEPH) is one of the main reasons of severe pulmonary hypertension and has significantly higher morbidity and mortality rates. The pathogenesis of the disease is characterized by the incomplete resolution of acute embolisms. The elevated inflammatory conditions after the acute embolism are one of the critical factors. Therefore, we aimed to investigate whether or not anakinra is an option for treating CTEPH in an animal model. We studied twenty-one rats in this study They were randomly divided into three groups containing seven animals: the control group: saline-treated control; the embolism group: CTEPH + normal saline; the anakinra group: CTEPH + anakinra. We have observed that the layers of the segmental arteries and the alveolar were normal in the control group. In the cardiac tissue it was observed that muscular tissues and connective tissue were normal in the right ventricle. In embolism group, we detected a widening of the alveolar septum, a surrounding the alveolar infiltrates and a thickening of the segmental arteries in the muscular layer and a hypertrophy in the right ventricle tissues. We have determined that the lung and cardiac tissue specimens in the anakinra group are similar to control group. We have showed that anakinra was useful option for the CTEPH model in rats. Anakinra may be considered as protective effect and the regression of the increased inflammation in CTEPH. The effectiveness of anakinra will continue to be subject to the further experimental and clinical studies.

Model of chronic thromboembolic pulmonary hypertension in rats, caused by repeated intravenous administration of biodegradable microspheres from sodium alginate

Introduction. Chronic thromboembolic pulmonary hypertension (CTEPH) is one of the most severe complications of pulmonary embolism (PE), characterized by poor prognosis and insuffcient effectiveness of standard treatment approaches. A small number of representative models of CTEPH make it diffcult to conduct preclinical studies of promising pharmacological substances.Objective – development and validation of the experimental model of CTEPH in rats by embolization of the distal branches of the pulmonary artery with biodegradable microspheres.Material and methods. Male Wistar rats were used for the experiments. Biodegradable microspheres (MS) based on sodium alginate and autologous blood clots (AT) were used as embolizing particles. The animals were divided into groups: control: saline solution was injected 4 times with an interval of 8 days into the tail vein; AT: according to the above protocol, 50 μL of AT was injected; MS was administered intravenously in a volume of 50 μl of MS according to two protocols: MS4: 4 times with an interval of 8 days; MS8: 8 times with an interval of 4 days. After 2 and 6 weeks after the last injection, a histological examination of the lungs was performed; after 6 weeks: echocardiographic study (TTE), right ventricular catheterization (RV) with measurement of right ventricular systolic pressure (RVSP), treadmill test, assessment of serum endothelin­1 levels by the immunoassay method.Results. During the experiments, the survival rate in the MS8 group was 50 %. In the other groups, there were no animal losses. According to the treadmill test 6 weeks after the modeling of PE, exercise tolerance was signifcantly reduced in the MC4 and MC8 groups compared with the control group. TTE data indicate a signifcant increase in the diameter of the pulmonary trunk and the right ventricular outflow tract in the MC8 compared with the control and AT. There were signifcant increase in RVSP and the level of endothelin­1 compared with the control only in the MS8. After 6 weeks, the index of hypertrophy of vessel wall of the pulmonary artery in the MC4 and MC8 was signifcantly higher compared with the control and AT groups.Conclusion. Based on the use of MS, administered under the MS 8 protocol, a new representative model of CTEPH has been created, which can be used to test promising pharmacological substances.

Right ventricular mitochondrial respiratory function in a piglet model of chronic pulmonary hypertension

ObjectiveWe aimed to assess the mitochondrial respiratory capacities in the right ventricle in the setting of ventricular remodeling induced by pressure overload. MethodsChronic thromboembolic pulmonary hypertension was induced in 8 piglets over a 12-week period (chronic thromboembolic pulmonary hypertension model). Right ventricular remodeling, right ventricular function, and mitochondrial respiratory function were assessed at 3, 6, and 12 weeks after induction of pulmonary hypertension and were compared with sham animals (n = 5). Right ventricular cardiomyocytes and mitochondrial structure were studied in transmission electronic microscopy after 12 weeks. ResultsAs of 3 weeks, chronic pressure overload induced right ventricular dilatation, right ventricular hypertrophy, and right ventricular dysfunction. Maladaptive remodeling in the chronic thromboembolic pulmonary hypertension model was confirmed by the decrease of right ventricular pulmonary artery coupling and right fractional area change. Mitochondrial functional assays in permeabilized right ventricular myocardial fibers revealed that oxidative phosphorylation capacities (complex I, complex II, and IV of the mitochondrial respiratory chain) were degraded. Furthermore, no change in substrate preference of mitochondria was found in the overloaded right ventricle. There was a good correlation between maximal mitochondrial oxygen consumption rate and right ventricular pulmonary artery coupling (Pearson coefficient r = 0.83). Transmission electronic microscopy analysis showed that the composition of cardiomyocytes was no different between the chronic thromboembolic pulmonary hypertension group and the sham group. However, mitochondrial structure anomalies were significantly increased in the chronic thromboembolic pulmonary hypertension group. ConclusionsMitochondrial respiratory function impairment is involved early in the development of right ventricular dysfunction in a piglet model of chronic thromboembolic pulmonary hypertension. Underlying mechanisms remain to be elucidated.

A Large Animal Model of Right Ventricular Failure due to Chronic Thromboembolic Pulmonary Hypertension: A Focus on Function.

Chronic thromboembolic pulmonary hypertension (CTEPH) is a debilitating disease that progresses to right ventricular (RV) failure and death if left untreated. Little is known regarding the progression of RV failure in this disease, greatly limiting effective prognoses, and therapeutic interventions. Large animal models enable the use of clinical techniques and technologies to assess progression and diagnose failure, but the existing large animal models of CTEPH have not been shown to replicate the functional consequences of the RV, i.e., RV failure. Here, we created a canine embolization model of CTEPH utilizing only microsphere injections, and we used a combination of right heart catheterization (RHC), echocardiography (echo), and magnetic resonance imaging (MRI) to quantify RV function. Over the course of several months, CTEPH led to a 6-fold increase in pulmonary vascular resistance (PVR) in four adult, male beagles. As evidenced by decreased cardiac index (0.12 ± 0.01 v. 0.07 ± 0.01 [L/(min*kg)]; p < 0.05), ejection fraction (0.48 ± 0.02 v. 0.31 ± 0.02; p < 0.05), and ventricular-vascular coupling ratio (0.95 ± 0.09 v. 0.45 ± 0.05; p < 0.05), as well as decreased tricuspid annular plane systolic excursion (TAPSE) (1.37 ± 0.06 v. 0.86 ± 0.05 [cm]; p < 0.05) and increased end-diastolic volume index (2.73 ± 0.06 v. 2.98 ± 0.02 [mL/kg]; p < 0.05), the model caused RV failure. The ability of this large animal CTEPH model to replicate the hemodynamic consequences of the human disease suggests that it could be utilized for future studies to gain insight into the pathophysiology of CTEPH development, following further optimization.

Autologous endothelial progenitor cell therapy improves right ventricular function in a model of chronic thromboembolic pulmonary hypertension

BackgroundRight ventricular (RV) failure is the main prognostic factor in pulmonary hypertension, and ventricular capillary density (CD) has been reported to be a marker of RV maladaptive remodeling and failure. Our aim was to determine whether right intracoronary endothelial progenitor cell (EPC) infusion can improve RV function and CD in a piglet model of chronic thromboembolic pulmonary hypertension (CTEPH). MethodsWe compared 3 groups: sham (n = 5), CTEPH (n = 6), and CTEPH with EPC infusion (CTEPH+EPC; n = 5). After EPC isolation from CTEPH+EPC piglet peripheral blood samples at 3 weeks, the CTEPH and sham groups underwent right intracoronary infusion of saline, and the CTEPH+EPC group received EPCs at 6 weeks. RV function, pulmonary hemodynamics, and myocardial morphometry were investigated in the animals at 10 weeks. ResultsAfter EPC administration, the RV fractional area change increased from 32.75% (interquartile range [IQR], 29.5%-36.5%) to 39% (IQR, 37.25%-46.50%; P = .030). The CTEPH+EPC piglets had reduced cardiomyocyte surface areas (from 298.3 μm2 [IQR, 277.4-335.3 μm2] to 234.6 μm2 (IQR, 211.1-264.7 μm2; P = .017), and increased CD31 expression (from 3.12 [IQR, 1.27-5.09] to 7.14 [IQR, 5.56-8.41; P = .017). EPCs were found in the RV free wall at 4 and 24 hours after injection but not 4 weeks later. ConclusionsIntracoronary infusion of EPC improved RV function and CD in a piglet model of CTEPH. This novel cell-based therapy might represent a promising RV-targeted treatment in patients with pulmonary hypertension.

The importance of capillary density–stroke work mismatch for right ventricular adaptation to chronic pressure overload

ObjectiveMechanisms of right ventricular (RV) adaptation to chronic pressure overload are not well understood. We hypothesized that a lower capillary density (CD) to stroke work ratio would be associated with more fibrosis and RV maladaptive remodeling. MethodsWe induced RV chronic pressure overload over a 20-week period in 2 piglet models of pulmonary hypertension; that is, a shunt model (n = 5) and a chronic thromboembolic pulmonary hypertension model (n = 5). We assessed hemodynamic parameters and RV remodeling as well as RV CD, fibrosis, and angiogenic factors expression. ResultsAlthough RV was similarly hypertrophied in both models, maladapted RV remodeling with impaired systolic function was only seen in chronic thromboembolic pulmonary hypertension group members who had lower CD (484 ± 99 vs 1213 ± 74 cap/mm2; P < .01), lower CD to stroke work ratio (0.29 ± 0.07 vs 0.82 ± 0.16; P = .02), higher myocardial fibrosis (15.4% ± 3.8% vs 8.0% ± 2.5%; P < .01), as well as a higher angiogenic and fibrosis factors expression. ConclusionsThe RV adaptive response to chronic pressure overload differs between 2 different piglet models of PH. Mismatch between angiogenesis and workload (CD to stroke work ratio) was associated with greater degree of myocardial fibrosis and RV dysfunction and could be a promising index of RV maladaptation. Further studies are needed to understand the underlying mechanisms.

Pulmonary arterial strain- and remodeling-induced stiffening are differentiated in a chronic model of pulmonary hypertension

Pulmonary hypertension (PH) is a debilitating vascular disease that leads to pulmonary artery (PA) stiffening, which is a predictor of patient mortality. During PH development, PA stiffening adversely affects right ventricular function. PA stiffening has been investigated through the arterial nonlinear elastic response during mechanical testing using a canine PH model. However, only circumferential properties were reported and in the absence of chronic PH-induced PA remodeling. Remodeling can alter arterial nonlinear elastic properties via chronic changes in extracellular matrix (ECM) content and geometry. Here, we used an established constitutive model to demonstrate and differentiate between strain-stiffening, which is due to nonlinear elasticity, and remodeling-induced stiffening, which is due to ECM and geometric changes, in a canine model of chronic thromboembolic PH (CTEPH). To do this, circumferential and axial tissue strips of large extralobar PAs from control and CTEPH tissues were tested in uniaxial tension, and data were fit to a phenomenological constitutive model. Strain-induced stiffening was evident from mechanical testing as nonlinear elasticity in both directions and computationally by a high correlation coefficient between the mechanical data and model (R2=0.89). Remodeling-induced stiffening was evident from a significant increase in the constitutive model stress parameter, which correlated with increased PA collagen content and decreased PA elastin content as measured histologically. The ability to differentiate between strain- and remodeling-induced stiffening in vivo may lead to tailored clinical treatments for PA stiffening in PH patients.

Expression of tissue factor and forkhead box transcription factor O-1 in a rat model for chronic thromboembolic pulmonary hypertension.

Few reports have examined tissue factor (TF) and forkhead box transcription factor O-1 (FoxO1) expression in chronic thromboembolic pulmonary hypertension (CTEPH) animal models. To investigate the role of TF and FoxO1 and their interactions during CTEPH pathogenesis in a rat model. Autologous blood clots were repeatedly injected into the pulmonary arteries through right jugular vein to induce a rat model of CTEPH. Hemodynamic parameters, histopathology, and TF and FoxO1expression levels were detected. The mean pulmonary arterial pressure (mPAP), pulmonary vascular resistance and vessel wall area/total area (WA/TA) ratio in the experiment group increased significantly than sham group (P<0.05). The cardiac output in the 1-, 2-, and 4-week groups decreased significantly (P<0.05) when compared to sham group. TF mRNA expression levels in the experiment group increased significantly than sham group (P<0.05). FoxO1 mRNA and protein expression levels were lower in the experiment group than sham group (P<0.05). The mPAP had a positive correlation with WA/TA ratio (r=0.45, P=0.01). TF mRNA expression had a positive correlation with WA/TA ratio (r=0.374, P=0.035) and a positive correlation with mPAP (r=0.48, P=0.005). FoxO1 mRNA expression had a negative correlation trend with the WA/TA ratio (r=-0.297, P=0.099) and a negative correlation trend with mPAP (r=-0.34, P=0.057). TF mRNA expression had a negative correlation with FoxO1 mRNA expression (r=-0.62, P<0.001). A rat model of CTEPH can be successfully established by the injection of autologous blood clots into the pulmonary artery. TF and FoxO1 may play a key role in vascular remodeling during CTEPH pathogenesis.

The role of tissue factor and autophagy in pulmonary vascular remodeling in a rat model for chronic thromboembolic pulmonary hypertension.

BackgroundFew reports have examined tissue factor (TF) and autophagy expression in chronic pulmonary thromboembolic hypertension (CTEPH) animal models. Objectives: To investigate the role of tissue factor (TF), autophagy and their interactions during chronic thromboembolic pulmonary hypertension (CTEPH) pathogenesis in a rat model.MethodsAutologous blood clots were repeatedly injected into the left jugular vein of rats with injecting endogenous fibrinolysis inhibitor tranexamic acid (TXA). Mean pulmonary arterial pressure (mPAP), histopathology and TF, Beclin-1 and microtubule-associated protein 1 light chain (LC3) expression levels were detected.ResultsThe mPAP and vessel wall area/total area (WA/TA) ratio in the experiment group increased significantly (P < 0.05). TF mRNA and protein expression levels in the experiment group increased significantly (P < 0.05). Beclin-1 and LC3B mRNA and protein expression levels were lower in the experiment group (P < 0.05). The mPAP had a positive correlation with WA/TA ratio (r = 0.955, P < 0.05). Beclin-1 and LC3B protein expression had a negative correlation with the WA/TA ratio (r = -0.963, P < 0.05, r = -0.965, P < 0.05, respectively). TF protein expression had a negative correlation with both Beclin-1 and LC3B protein expression (r = -0.995, P <0.05, r = -0972, P < 0.05, respectively).ConclusionsA rat model of CTEPH can be established by repeatedly introducing autologous blood clots into the pulmonary artery with injecting TXA. TF and autophagy may play a key role during CTEPH pathogenesis, especially in vascular remodeling.