Intrapulmonary Arteries Research Articles

The role of adipose tissue cell elements in the regulation of the nitroxidergic system and possible ways of pharmacological modulation

Endothelial dysfunction is characterized by a decrease in the bioavailability of the vasodilator – nitric oxide (NO), and an increase in the level of vasoconstrictor substances. This imbalance leads to vasoconstriction, leukocyte attachment and inflammatory reactions in the vascular wall, atherosclerosis and thrombosis. The aim: to evaluate the role of adipose tissue elements in the regulation of parameters of the nitroxidergic system under hypoxia conditions. Materials and methods. The studies were carried out on 30 adult white male Wistar rats. All animals were randomly assigned and divided into groups: a control group (15 rats), type 2 diabetes mellitus (T2DM) was induced in the animals of the second group (15 rats). Isolated fragments of the popliteal arteries (PA) and intrapulmonary artery (IPA) were cleared of perivascular adipose tissue (PVAT-) or left uncleaned (PVAT+) and cut into rings. The simulation of acute hypoxia with further study of medical agents were performed. Results. The PA and IPA with PVAT responded to acute hypoxia with vasoconstriction – an increase in the amplitude of contraction in the first and second phases, and after removing PVAT, they responded with a decrease in the maximum amplitude of contraction by 3.4 times in the 1st phase and an increase in amplitude by 1.8 times in the 2nd phase. Perfusion with Angiolin reduced 2nd phase of HV of the PA and IPA. Adding a combination of Thiotriazoline and L-arginine (1:4) to a solution for perfusion of fragments of arteries of animals with T2DM, causes a significant increase in constrictor reactions in both the 1st and 2nd phases of HV, regardless of presence of perivascular adipose tissue. Conclusions. The presence of PVAT affects the HV of arteries, both in normal and in T2DM. The possibilities of ways of pharmacological modulation of the nitroxidergic system depending on the state of PVAT were determined.

Ethanol-induced lung and cardiac right ventricular inflammation and remodeling underlie progression to pulmonary arterial hypertension.

Current research on ethanol-induced cardiovascular anomalies has focused on left ventricular (LV) function and blood pressure. To extend this area of research, we sought to determine whether ethanol-induced alterations in the structure and function of the right cardiac ventricle (RV) and pulmonary artery (PA) lead to pulmonary arterial hypertension (PAH). Two groups of male Sprague-Dawley rats received a balanced liquid diet containing 5% ethanol (w/v) or a pair-fed isocaloric liquid diet for 8 weeks. Weekly echocardiography was conducted to evaluate cardiopulmonary function, and lung and RV tissues were collected for exvivo histological and molecular studies. The ethanol-treated rats exhibited: (1) Elevated mean pulmonary arterial pressure and decreased pulmonary artery acceleration time/ejection time; (2) Pulmonary vascular remodeling comprising intrapulmonary artery medial layer thickening; and (3) RV hypertrophy along with increased RV/LV + septum, RV diameter, RV cardiomyocyte cross-sectional area, and LV mass/body weight ratio. These responses were associated with increased lung and RV pro-inflammatory markers, endothelin-1 (ET-1), TNF-α, and IL-6 levels and higher ET-1,ET-1 type A/B receptor ratio, and downregulation of the cytoprotective protein, bone morphogenetic protein receptor 2 (BMPR2), in the lungs. These findings show that moderate ethanol-induced cardiopulmonary changes underlie progression to PAH via an upregulated proinflammatory ET1-TNFα-IL6 pathway and suppression of the anti-inflammatory BMPR2.

Pulmonary vascular reactivity in growth restricted fetuses using computational modelling and machine learning analysis of fetal Doppler waveforms

The aim of this study was to investigate the pulmonary vasculature in baseline conditions and after maternal hyperoxygenation in growth restricted fetuses (FGR). A prospective cohort study of singleton pregnancies including 97 FGR and 111 normally grown fetuses was carried out. Ultrasound Doppler of the pulmonary vessels was obtained at 24–37 weeks of gestation and data were acquired before and after oxygen administration. After, Machine Learning (ML) and a computational model were used on the Doppler waveforms to classify individuals and estimate pulmonary vascular resistance (PVR). Our results showed lower mean velocity time integral (VTI) in the main pulmonary and intrapulmonary arteries in baseline conditions in FGR individuals. Delta changes of the main pulmonary artery VTI and intrapulmonary artery pulsatility index before and after hyperoxygenation were significantly greater in FGR when compared with controls. Also, ML identified two clusters: A (including 66% controls and 34% FGR) with similar Doppler traces over time and B (including 33% controls and 67% FGR) with changes after hyperoxygenation. The computational model estimated the ratio of PVR before and after maternal hyperoxygenation which was closer to 1 in cluster A (cluster A 0.98 ± 0.33 vs cluster B 0.78 ± 0.28, p = 0.0156). Doppler ultrasound allows the detection of significant changes in pulmonary vasculature in most FGR at baseline, and distinct responses to hyperoxygenation. Future studies are warranted to assess its potential applicability in the clinical management of FGR.

Pathological features of primary cardiac myxoid tumour in dogs: a review of 11 cases (2002–2022)

This report documents the pathological features of primary cardiac myxoid tumour (MT) in 11 dogs. Macroscopically, all the tumours were located in the tricuspid valve (TV), its septal leaflet being predominantly affected. Therefore, it appears that the TV is the most common site of occurrence for cardiac MT in dogs. Two gross anatomical types of canine valvular MT were evident. Seven of the 11 tumours were round or oval with a smooth or gently lobulated and glistening surface, while the other four were gelatinous, multilobulated and polypoid, with an irregular surface. Microscopically, in nine cases the tumours had an abundant myxoid matrix within which elongated spindle-shaped cells with no remarkable cytological atypia were sparsely embedded, suggesting a benign character (ie, myxoma). In the other two cases the tumours consisted of variably dense, haphazardly arranged, interlacing streams of anaplastic spindle-shaped or polygonal cells containing many mitotic figures, indicative of a malignant form of myxoma (ie, myxosarcoma). Isolated or clustered collections of myxoma cells (eg, cords, rings, syncytia) characteristic of human atrial myxoma were only rarely evident or lacking in all 11 cases, indicating that rarity or absence of such structural features may be specific to valvular MTs. Immunohistochemical findings were indicative of smooth muscle differentiation of the neoplastic cells. Tumour embolization to the intrapulmonary arteries and/or tumour implantation on the endocardium of the right heart chambers was evident only in the four cases of irregular-surfaced MT.

The small-molecule formyl peptide receptor biased agonist, compound 17b, is a vasodilator and anti-inflammatory in mouse precision-cut lung slices.

Pulmonary arterial hypertension (PAH), a rare fatal disorder characterised by inflammation, vascular remodelling and vasoconstriction. Current vasodilator therapies reduce pulmonary arterial pressure but not mortality. The G-protein coupled formyl peptide receptors (FPRs) mediates vasodilatation and resolution of inflammation, actions possibly beneficial in PAH. We investigated dilator and anti-inflammatory effects of the FPR biased agonist compound 17b in pulmonary vasculature using mouse precision-cut lung slices (PCLS). PCLS from 8-week-old male and female C57BL/6 mice, intrapulmonary arteries were pre-contracted with 5-HT for concentration-response curves to compound 17b and 43, and standard-of-care drugs, sildenafil, iloprost and riociguat. Compound 17b-mediated relaxation was assessed with FPR antagonists or inhibitors and in PCLS treated with TNF-α or LPS. Cytokine release from TNF-α- or LPS-treated PCLS ± compound 17b was measured. Compound 17b elicited concentration-dependent vasodilation, with potencies of iloprost > compound 17b = riociguat > compound 43 = sildenafil. Compound 17b was inhibited by the FPR1 antagonist cyclosporin H but not by soluble guanylate cyclase, nitric oxide synthase or cyclooxygenase inhibitors. Under inflammatory conditions, the efficacy and potency of compound 17b were maintained, while iloprost and sildenafil were less effective. Additionally, compound 17b inhibited secretion of PAH-relevant cytokines via FPR2. Vasodilation to compound 17b but not standard-of-care vasodilators, is maintained under inflammatory conditions, with additional inhibition of PAH-relevant cytokine release. This provides the first evidence that targeting FPR, with biased agonist, simultaneously targets vascular function and inflammation, supporting the development of FPR-based pharmacotherapy to treat PAH. This article is part of a themed issue Therapeutic Targeting of G Protein-Coupled Receptors: hot topics from the Australasian Society of Clinical and Experimental Pharmacologists and Toxicologists 2021 Virtual Annual Scientific Meeting. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v181.14/issuetoc.

DOTATOC PET/CT imaging of a typical carcinoid tumor in a human ex-vivo perfused lung lobe.

The use of Isolated lung perfusion (ILP), combined with medical imaging modalities such as positron emission tomography-computed tomography (PET/CT), provides real-time visualization of tumors in ventilated and perfused vital lung tissue. This experiment intends to show the feasibility and benefits of using ILP combined with PET/CT imaging. Following lung surgery on a 49-year-old male, his left lower lobectomy specimen, which held a typical carcinoid tumor, was preserved on normothermic ILP. Gallium-68-Edotreotide ([68Ga]-DOTATOC) was administered into the ILP circuit, and dynamic emission data from PET/CT was acquired. ILP was carried out for 120 minutes. Near physiologic gas exchange and glucose metabolism were preserved throughout the experiment. The time activity curves (TAC) of 5 different volumes of interest (VOI) showed notable differences in tracer uptake over time. The peripheral area of the carcinoid exhibited delayed but high somatostatin receptor agonist uptake compared to the surrounding parenchyma and the intrapulmonary artery. However, the central area of the carcinoid showed very low [68Ga]-DOTATOC uptake. This experiment demonstrates the potential of ILP combined with PET/CT for kinetic modeling in experimental nuclear medicine imaging. By providing visualization of tracer uptake in perfused lung tissue, this model could potentially improve our understanding of tumor physiology and molecular imaging.

Intrapulmonary arterial contraction assay reveals region-specific deregulation of vasoreactivity to lung injuries.

Intrapulmonary arteries located in the proximal lung differ from those in the distal lung in size, cellular composition, and the surrounding microenvironment. However, whether these structural variations lead to region-specific regulation of vasoreactivity in homeostasis and following injury is unknown. Herein, we employ a two-step method of precision-cut lung slice (PCLS) preparation, which maintains almost intact intrapulmonary arteries, to assess contractile and relaxation responses of proximal preacinar arteries (PaAs) and distal intraacinar arteries (IaAs) in mice. We found that PaAs exhibited robust vasoconstriction in response to contractile agonists and significant nitric oxide (NO)-induced vasodilation. In comparison, IaAs were less contractile and displayed a greater relaxation response to NO. Furthermore, in a mouse model of pulmonary arterial hypertension (PAH) induced by chronic exposure to ovalbumin (OVA) allergen and hypoxia (OVA-HX), IaAs demonstrated a reduced vasocontraction despite vascular wall thickening with the emergence of new αSMA+ cells coexpressing markers of pericytes. In contrast, PaAs became hypercontractile and less responsive to NO. The reduction in relaxation of PaAs was associated with decreased expression of protein kinase G, a key component of the NO pathway, following chronic OVA-HX exposure. Taken together, the PCLS prepared using the modified preparation method enables functional evaluation of pulmonary arteries in different anatomical locations and reveals region-specific mechanisms underlying the pathophysiology of PAH in a mouse model.NEW & NOTEWORTHY Utilizing mouse precision-cut lung slices with preserved intrapulmonary vessels, we demonstrated a location-dependent structural and contractile regulation of pulmonary arteries in health and on noxious stimulations. For instance, chronic ovalbumin and hypoxic exposure increased pulmonary arterial pressure (PAH) by remodeling intraacinar arterioles to reduce vascular wall compliance while enhancing vasoconstriction in proximal preacinar arteries. These findings suggest region-specific mechanisms and therapeutic targets for pulmonary vascular diseases such as PAH.

Acid-sensing ion channel 1a regulates mitochondrial membrane potential and pulmonary arterial smooth muscle cell apoptosis

Acid-sensing ion channel 1a (ASIC1a) is a proton-gated cation channel that contributes to the development of chronic hypoxia (CH)-induced pulmonary hypertension (PH). Our prior studies demonstrate that ASIC1a is expressed in pulmonary arterial smooth muscle cells (PASMCs) and contributes to plasmalemmal membrane potential ( E m ) depolarization and augmented intracellular Ca 2+ signaling leading to enhanced vasoconstrictor reactivity and vascular remodeling. During PH, mitochondrial dysfunction associated with mitochondrial membrane potential (Δψm) hyperpolarization can inhibit opening of the mitochondrial permeability transition pore (mPTP), thereby repressing apoptosis and promoting the hyperproliferative and apoptosis-resistant PASMC phenotype. Recent evidence suggests that ASIC1a is localized to mitochondria and regulates the mPTP. However, it is unknown if mitochondrial-localized ASIC1a (mtASIC1a) contributes to the pathogenesis of PH. Therefore, the overall objective of this study is to determine the functional role of mtASIC1a in the apoptosis-resistant PASMC phenotype associated with PH. We hypothesized that CH causes Δψm hyperpolarization and apoptosis-resistance in PASMCs due to loss of mtASIC1a localization. To test this hypothesis, we housed Asic1a knockout ( Asic1a -/- ) and wild-type ( Asic1a +/+ ) mice in a hypobaric chamber (barometric pressure: ~380 mmHg) for 4 weeks to induce PH. Following CH, we found increased expression of ASIC1a at the plasma membrane and decreased expression in mitochondria, which were associated with PASMC E m depolarization and Δψm hyperpolarization. Using tetramethylrhodamine (TMRE) fluorescence to measure Δψm, we found that TMRE fluorescence was significantly increased (p=0.02) in PASMCs from normoxic Asic1a -/- mice compared to Asic1a +/+ mice, indicating that deletion of Asic1a leads to Δψm hyperpolarization. We utilized a lentiviral vector encoding mitochondrial-targeted human ASIC1a (mtASIC1a LV) to overexpress mtASIC1a in PASMCs (p=0.003) and found that transduction with mtASIC1a LV in PASMCs from Asic1a -/- mice significantly decreased TMRE fluorescence (p=0.004), further supporting the functional role of mtASIC1a to regulate Δψm. To assess apoptosis, we measured levels of active caspase-3 by western blot analysis in intrapulmonary arteries (PAs) from Asic1a +/+ and Asic1a -/- mice following CH. CH decreased levels of active caspase-3 in PAs from Asic1a +/+ (p=0.032), but not Asic1a -/- mice. Under normoxic conditions, active caspase-3 was lower in PAs from Asic1a -/- compared to Asic1a +/+ mice (p=0.044). These data suggest mtASIC1a plays an important role in the regulation of Δψm and that the loss of mtASIC1a contributes to Δψm hyperpolarization and apoptosis resistance in PASMCs following CH. Future studies will examine the contributions of mtASIC1a to the pathogenesis of PH through the regulation of mitochondrial reactive oxygen species, mitochondrial Ca 2+ levels, mPTP sensitivity, and mitochondrial dynamics. NIH R01 HL111084 to N.L. Jernigan and NIH T32 HL007736 to T.C. Resta This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Characterisation of P2Y receptor subtypes mediating vasodilation and vasoconstriction of rat pulmonary artery using selective antagonists

Pulmonary vascular tone is modulated by nucleotides, but which P2 receptors mediate these actions is largely unclear. The aim of this study, therefore, was to use subtype-selective antagonists to determine the roles of individual P2Y receptor subtypes in nucleotide-evoked pulmonary vasodilation and vasoconstriction. Isometric tension was recorded from rat intrapulmonary artery rings (i.d. 200–500 µm) mounted on a wire myograph. Nucleotides evoked concentration- and endothelium-dependent vasodilation of precontracted tissues, but the concentration–response curves were shallow and did not reach a plateau. The selective P2Y2 antagonist, AR-C118925XX, inhibited uridine 5′-triphosphate (UTP)- but not adenosine 5′-triphosphate (ATP)-evoked relaxation, whereas the P2Y6 receptor antagonist, MRS2578, had no effect on UTP but inhibited relaxation elicited by uridine 5′-diphosphate (UDP). ATP-evoked relaxations were unaffected by the P2Y1 receptor antagonist, MRS2179, which substantially inhibited responses to adenosine 5′-diphosphate (ADP), and by the P2Y12/13 receptor antagonist, cangrelor, which potentiated responses to ADP. Both agonists were unaffected by CGS1593, an adenosine receptor antagonist. Finally, AR-C118925XX had no effect on vasoconstriction elicited by UTP or ATP at resting tone, although P2Y2 receptor mRNA was extracted from endothelium-denuded tissues using reverse transcription polymerase chain reaction with specific oligonucleotide primers. In conclusion, UTP elicits pulmonary vasodilation via P2Y2 receptors, whereas UDP acts at P2Y6 and ADP at P2Y1 receptors, respectively. How ATP induces vasodilation is unclear, but it does not involve P2Y1, P2Y2, P2Y12, P2Y13, or adenosine receptors. UTP- and ATP-evoked vasoconstriction was not mediated by P2Y2 receptors. Thus, this study advances our understanding of how nucleotides modulate pulmonary vascular tone.

The Role and Clinical Value of Optimized Fetal Main Pulmonary Artery Doppler Parameters in the Diagnosis and Prognosis Monitoring of Neonatal Respiratory Distress Syndrome.

Objective A retrospective study was conducted to explore the role and clinical value of optimized fetal main pulmonary artery Doppler parameters in the diagnosis and prognosis monitoring of neonatal respiratory distress syndrome (NRDS). Methods Pregnant women admitted to our hospital for delivery from March 2019 to June 2021 were included in the study. There were 372 singleton pregnant women who voluntarily underwent ultrasound and Doppler examinations with informed consent for the study. The AT/ET ratio of the middle segment of the main pulmonary artery, the middle part of the left (or right) pulmonary artery, and the proximal segment of the intrapulmonary artery was measured by ultrasonic Doppler. According to the outcome of the newborn, it was divided into NRDS group (n =90) and non-NRDS group (n =282) to compare the differences of fetal AT/ET ratio at three Doppler sampling locations and the differences of fetal AT/ET ratio in different gestational weeks. Using Pearson's correlation analysis, the correlation among the Doppler parameters of fetal main pulmonary artery, the ratio of AT and AT/ET, gestational weeks, and the efficiency of prenatal prediction of NRDS was evaluated by receiver-operating characteristic (ROC). In addition, the diagnostic efficiency of fetal main pulmonary artery Doppler parameter AT/ET ratio and amniotic fluid method for prenatal prediction of NRDS was compared. Results Compared with the non-NRDS group, the gestational age, gestational age at delivery, and birth weight in the NRDS group were significantly lower than those in the non-NRDS group. The 1 min Apgar score of newborns was less than 7, and the proportion of newborns transferred to NICU was significantly higher than that in the control group. Compared with non-NRDS group, AT was significantly shortened, AT/ET ratio was significantly decreased, and ET was significantly prolonged in NRDS group. The fetal aortic blood flow parameters AT, ET, and AT/ET ratio in different gestational weeks were significantly different among the three groups. The AT/ET ratio in the middle segment of the aorta in NRDS group was lower than that in non-NRDS group. The ratio of AT/ET was still correlated with the occurrence of postpartum NRDS. The AUC value of NRDS predicted by AT/ET ratio was 0.984. In the combined determination of amniotic fluid Lh and S ratio and PG, the diagnostic accuracy of prenatal prediction of neonatal respiratory distress syndrome was 100.005%, 98.43%, and 95.56%, respectively. The accuracy of fetal aortic AT/ET ratio in prenatal prediction of neonatal respiratory distress syndrome was 100.005%, 95.28%, and 95.57%, respectively. The order of Kappa coefficient from high to low was amniotic fluid PG/L/S ratio, fetal main pulmonary artery blood flow parameter AT/ET ratio, amniotic fluid L/S ratio, OD650, amniotic fluid PG, and foam test. Conclusion Fetal main pulmonary artery Doppler AT/ET ratio may be a very potential parameter for prenatal noninvasive prediction of NRDS. The fetal main pulmonary artery Doppler flow spectrum may be a very potential alternative to amniocentesis to evaluate FLM.

Piezo1 Channel Activation Reverses Pulmonary Artery Vasoconstriction in an Early Rat Model of Pulmonary Hypertension: The Role of Ca2+ Influx and Akt-eNOS Pathway.

In intrapulmonary arteries (IPAs), mechanical forces due to blood flow control vessel tone, and these forces change during pulmonary hypertension (PH). Piezo1, a stretch-activated calcium channel, is a sensor of mechanical stress present in both endothelial cells (ECs) and smooth muscle cells (SMCs). The present study investigated the role of Piezo1 on IPA in the chronic hypoxia model of PH. Rats were raised in chronically hypoxic conditions for 1 (1W-CH, early stage) or 3 weeks (3W-CH, late-stage) of PH or in normoxic conditions (Nx). Immunofluorescence labeling and patch-clamping revealed the presence of Piezo1 in both ECs and SMCs. The Piezo1 agonist, Yoda1, induced an IPA contraction in Nx and 3W-CH. Conversely, Yoda1 induced an endothelial nitric oxide (eNOS) dependent relaxation in 1W-CH. In ECs, the Yoda1-mediated intracellular calcium concentration ([Ca2+]i) increase was greater in 1W-CH as compared to Nx. Yoda1 induced an EC hyperpolarization in 1W-CH. The eNOS levels were increased in 1W-CH IPA compared to Nx or 3W-CH PH and Yoda1 activated phosphorylation of Akt (Ser473) and eNOS (Ser1177). Thus, we demonstrated that endothelial Piezo1 contributes to intrapulmonary vascular relaxation by controlling endothelial [Ca2+]i, endothelial-dependent hyperpolarization, and Akt-eNOS pathway activation in the early stage of PH.

Isolation of Intrapulmonary Artery and Smooth Muscle Cells to Investigate Vascular Responses.

The intrapulmonary artery (IPA) and vascular smooth muscle cells (VSMCs) isolated from rat lungs can be used to study the underlying mechanisms of vasoconstriction and vasorelaxation. After isolating the IPA and VSMCs, the characteristics of vascular responses in physiological and pathological conditions can be assessed in the absence of extrinsic factors such as nerve signals, hormones, cytokines, etc. Thus, the IPA and VSMCs serve as excellent models for studying vascular physiology/pathophysiology, along with various experimental investigations, such as modulation by pharmacological agents, patch-clamp electrophysiological analysis, calcium imaging, etc. Here, we have used a technique for isolating the IPA to investigate vascular responses in an organ bath setup. IPA segments were mounted on the organ bath chamber via intraluminal wires and stimulated by various pharmacological agents. The changes in IPA vascular tone (i.e., vasoconstriction and vasorelaxation), were recorded using an isometric force transducer and physiological data analysis software program. We implemented several experimental protocols, which can be adapted to investigate the mechanisms of vasorelaxation/vasoconstriction for studying the pharmacological activities of phytochemical or synthetic drugs. The protocols can also be used to evaluate drugs' roles in modulating various diseases, including pulmonary arterial hypertension. The IPA model allows us to investigate the concentration-response curve, which is crucial in assessing drugs' pharmacodynamic parameters.

Isolation of Intrapulmonary Artery and Smooth Muscle Cells to Investigate Vascular Responses

Isolation of Intrapulmonary Artery and Smooth Muscle Cells to Investigate Vascular Responses

Targeting Piezo1 channels to reverse pulmonary artery vasoconstriction in rat model of pulmonary hypertension

Piezo1, a stretch-activated calcium permeable channel is a sensor of mechanical stress present in endothelial cells (ECs) and vascular smooth muscle cells (SMCs). Our previous studies have shown that endothelial Piezo1 contributes to intrapulmonary artery (IPA) relaxation in mice. The present study was undertaken to investigate the implication of Piezo1 in both rat ECs and SMCs in pulmonary hypertension (PH). Chronically hypoxic rats, an animal model of PH, were exposed to chronic hypoxia (CH, −50% O 2 for 1 week), whereas normoxic rats were housed in normoxic condition (NX). NX or CH IPA tone variation induced by YODA1 was quantified by myograph. Intracellular calcium concentration ([Ca2+]i) was determined with Cal520-AM probes. Protein levels of endothelial nitric oxide synthase (eNOS), phosphorylated eNOS (peNOS), Akt and pAkt were quantified by Western blot analysis. Immunofluorescence labeling and patch-clamp revealed the presence of Piezo1 in both ECs and SMCs. In IPAs, YODA1 induced a contraction in NX condition. By contrast, YODA1 induced an endothelium and nitric oxide-dependent relaxation in CH IPAs. Activation of Piezo1 channels by its agonist YODA1 increased [Ca2+]i. However, the [Ca2+]i increase in ECs was greater in CH as compared to NX condition. In addition, CH condition increased level of eNOS proteins as compared to NX condition and YODA1 induced a phosphorylation of Akt (Ser 473) and eNOS (Ser 1177) in NX and CH conditions. The present study thus demonstrates that endothelial Piezo1 contributes to intrapulmonary vascular relaxation by controlling endothelial [Ca2+]i and Akt-eNOS- pathway activation that could counteract PH.

Utilizing the Precision-Cut Lung Slice to Study the Contractile Regulation of Airway and Intrapulmonary Arterial Smooth Muscle.

Smooth muscle cells (SMC) mediate the contraction of the airway and the intrapulmonary artery to modify airflow resistance and pulmonary circulation, respectively, hence playing a critical role in the homeostasis of the pulmonary system. Deregulation of SMC contractility contributes to several pulmonary diseases, including asthma and pulmonary hypertension. However, due to limited tissue access and a lack of culture systems to maintain in vivo SMC phenotypes, molecular mechanisms underlying the deregulated SMC contractility in these diseases remain fully identified. The precision-cut lung slice (PCLS) offers an ex vivo model that circumvents these technical difficulties. As a live, thin lung tissue section, the PCLS retains SMC in natural surroundings and allows in situ tracking of SMC contraction and intracellular Ca2+ signaling that regulates SMC contractility. Here, a detailed mouse PCLS preparation protocol is provided, which preserves intact airways and intrapulmonary arteries. This protocol involves two essential steps before subjecting the lung lobe to slicing: inflating the airway with low-melting-point agarose through the trachea and infilling pulmonary vessels with gelatin through the right ventricle. The PCLS prepared using this protocol can be used for bioassays to evaluate Ca2+-mediated contractile regulation of SMC in both the airway and the intrapulmonary arterial compartments. When applied to mouse models of respiratory diseases, this protocol enables the functional investigation of SMC, thereby providing insight into the underlying mechanism of SMC contractility deregulation in diseases.

68 Ga-FAPI-04 Versus 18 F-FDG PET/CT in a Case With Intimal Sarcoma of the Pulmonary Artery.

Intimal sarcoma of the pulmonary artery is a rare entity, mimicking acute or chronic pulmonary thromboembolism. We describe 68 Ga-FAPI-04 and 18 F-FDG PET/CT findings in a case with intimal sarcoma of the pulmonary artery. The intrapulmonary artery tumor showed inhomogeneously increased FDG and FAPI uptake. Compared with FDG PET/CT, FAPI PET/CT more clearly delineated the tumor partially because of a very low background FAPI activity in the walls of the main pulmonary artery and right ventricle, which showed diffuse FDG uptake.

Kv7 Channels in Cyclic-Nucleotide Dependent Relaxation of Rat Intra-Pulmonary Artery.

Pulmonary hypertension is treated with drugs that stimulate cGMP or cAMP signalling. Both nucleotides can activate Kv7 channels, leading to smooth muscle hyperpolarisation, reduced Ca2+ influx and relaxation. Kv7 activation by cGMP contributes to the pulmonary vasodilator action of nitric oxide, but its contribution when dilation is evoked by the atrial natriuretic peptide (ANP) sensitive guanylate cyclase, or cAMP, is unknown. Small vessel myography was used to investigate the ability of Kv7 channel blockers to interfere with pulmonary artery relaxation when cyclic nucleotide pathways were stimulated in different ways. The pan-Kv7 blockers, linopirdine and XE991, caused substantial inhibition of relaxation evoked by NO donors and ANP, as well as endothelium-dependent dilators, the guanylate cyclase stimulator, riociguat, and the phosphodiesterase-5 inhibitor, sildenafil. Maximum relaxation was reduced without a change in sensitivity. The blockers had relatively little effect on cAMP-mediated relaxation evoked by forskolin, isoprenaline or treprostinil. The Kv7.1-selective blocker, HMR1556, had no effect on cGMP or cAMP-dependent relaxation. Western blot analysis demonstrated the presence of Kv7.1 and Kv7.4 proteins, while selective activators of Kv7.1 and Kv7.4 homomeric channels, but not Kv7.5, caused pulmonary artery relaxation. It is concluded that Kv7.4 channels contribute to endothelium-dependent dilation and the effects of drugs that act by stimulating cGMP, but not cAMP, signalling.

Sodium tanshinone IIA sulfonate enhances the BMP9-BMPR2-Smad1/5/9 signaling pathway in rat pulmonary microvascular endothelial cells and human embryonic stem cell–derived endothelial cells

BackgroundRecent studies have demonstrated the beneficial effects of STS in treating pulmonary hypertension by inhibiting the pulmonary vascular remodeling and suppressing the abnormally elevated proliferation and migration of PASMCs. However, the roles of STS on pulmonary vascular endothelium remain largely known. MethodsIn this study, we investigated the effects and mechanisms of STS on pulmonary vascular endothelial dysfunction by using a chronic hypoxia-induced pulmonary hypertension (HPH) rat model, as well as in primarily cultured rat PMVECs and human ESC-ECs cell models. ResultsFirstly, a 21-day treatment of STS significantly prevents the disease development of HPH by normalizing the right ventricular systolic pressure and right ventricular hypertrophy, improving the cardiac output. Then, STS treatment markedly inhibits the hypoxia-induced medial wall thickening of the distal intrapulmonary arteries. Notably, STS significantly inhibits the hypoxia-induced apoptosis in both the pulmonary endothelium of HPH rats and primarily cultured PMVECs, through the stabilization of BMPR2 protein and protection of the diminished BMP9-BMPR2-Smad1/5/9 signaling pathway. In mechanism, STS treatment retrieves the hypoxic downregulation of BMPR2 by stabilizing the BMPR2 protein, inhibiting the BMPR2 protein degradation via lysosome system, and promoting the plasma membrane localization of BMPR2, all of which together reinforcing the BMP9-induced signaling transduction in both PMVECs and human ESC-ECs. However, these effects are absent in hESC-ECs expressing heterozygous dysfunctional BMPR2 protein (BMPR2+/R899X). ConclusionSTS may exert anti-apoptotic roles, at least partially, via induction of the BMP9-BMPR2-Smad1/5/9 signaling transduction in pulmonary endothelium and PMVECs.

Targeting Piezo1 channels to reverse pulmonary artery vasoconstriction in rat model of pulmonary hypertension

Piezo1, a stretch-activated calcium permeable channel is a sensor of mechanical stress present in endothelial cells (ECs) and vascular smooth muscle cells (SMCs). Our previous studies have shown that endothelial Piezo1 contributes to intrapulmonary artery (IPA) relaxation in mice. The present study was undertaken to investigate the implication of Piezo1 in both rat ECs and SMCs in pulmonary hypertension (PH). Chronically hypoxic rats, an animal model of PH, were exposed to chronic hypoxia (CH, −50% O2 for 1 weeks), whereas normoxic rats were housed in normoxic condition (NX). NX or CH IPA tone variation induced by YODA1 was quantified by myograph. Intracellular calcium concentration ([Ca2+]i) was determined with Cal520-AM probes. Protein levels of endothelial nitric oxide synthase (eNOS), phosphorylated eNOS (peNOS), Akt and pAkt were quantified by Western blot analysis. Immunofluorescence labeling and patch-clamp revealed the presence of Piezo1 in both ECs and SMCs. In IPAs, YODA1 induced a contraction in NX condition. By contrast, YODA1 induced an endothelium and nitric oxide-dependent relaxation in CH IPAs. Activation of Piezo1 channels by its agonist YODA1 increased [Ca2+]i. However, the [Ca2+]i increase in ECs was greater in CH as compared to NX condition. In addition, CH condition increased level of eNOS proteins as compared to NX condition and YODA1 induced a phosphorylation of Akt (Ser 473) and eNOS (Ser 1177) in NX and CH conditions. The present study thus demonstrates that endothelial Piezo1 contributes to intrapulmonary vascular relaxation by controlling endothelial [Ca2+]i and Akt-eNOS- pathway activation that could counteract PH.

Intrapulmonary artery septation for pulmonary artery growth in functionally univentricular hearts.

The pulmonary artery anatomy is one of the important determinants of Fontan completion and its outcomes in patients with functionally univentricular hearts. Despite several modifications to Fontan operation, unbalanced pulmonary arteries pose a significant challenge in this subset of patients. Intrapulmonary artery septation is a technique, developed to rehabilitate the unilateral hypoplastic pulmonary artery following various initial palliative procedures in functionally univentricular hearts. Literature is sparse on this technique. This article reviews the basic concepts and literature on intrapulmonary artery septation. We also describe our experience with modified intrapulmonary artery septation.