Recombinant thrombomodulin attenuates hyper-inflammation and glycocalyx damage in a murine model of Streptococcus pneumoniae-induced sepsis.

PURPOSE: The anticoagulant agent recombinant thrombomodulin (rTM) activates protein C to prevent excessive coagulation and also possibly regulates hyper-inflammation via neutralization of high-mobility-group B1 (HMG-B1). The glycocalyx layer in endothelial cells also plays a pivotal role in preventing septic shock-associated hyperpermeability. The present study examined the effect of rTM in a murine model of Streptococcus pneumoniae-induced sepsis. METHODS: Male C57BL/6N mice were injected intratracheally via midline cervical incision with 2 × 107 CFU of S. pneumoniae (capsular subtype 19A). Control mice were sham-treated identically but injected with saline. rTM (10 mg/kg) was injected intraperitoneally 3 h after septic insult. Blood concentrations of soluble inflammatory mediators (interleukin [IL]-1ß, IL-6, IL-10, and tumor necrosis factor [TNF]-α) were determined using a microarray immunoassay. Serum concentrations of HMG-B1 and syndecan-1, as a parameter of glycocalyx damage, were determined by enzyme-linked immunosorbent assay. The glycocalyx was also evaluated with electron microscopy. The lungs were removed, and digested to cells, which were then stained with a mixture of fluorophore-conjugated antibodies. Anti-mouse primary antibodies included PE-Cy7-conjugated anti-CD31, AlexaFluor 700-conjugated anti-CD45, PerCP-Cy5.5-conjugated anti-CD326, APC-conjugated anti-TNF-α, PE-conjugated anti-IL-6, and PE-conjugated anti-IL-10. A total of 1 × 106 cells per sample were analyzed, and 2 × 105 events were recorded by flow cytometry, and parameters were compared with/without rTM treatment. RESULTS: The blood concentration of TNF-α was significantly reduced 24 h after intratracheal injection in S. pneumoniae-challenged mice treated with rTM (P = 0.016). Levels of IL-10 in the lung endothelium of rTM-treated S. pneumoniae-challenged mice increased significantly 12 h after intratracheal injection (P = 0.03). Intriguingly, serum HMGB-1 and syndecan-1 levels decreased significantly (P = 0.010 and 0.015, respectively) in rTM-treated mice 24 h after intratracheal injection of S. pneumoniae. Electron microscopy indicated that rTM treatment preserved the morphology of the glycocalyx layer in septic mice. CONCLUSIONS: These data suggest that rTM modulates local inflammation in the lung endothelium, thus diminishing systemic inflammation, i.e., hypercytokinemia. Furthermore, rTM treatment reduced serum syndecan-1 levels, thus preventing glycocalyx damage. The use of rTM to treat sepsis caused by bacterial pneumonia could therefore help prevent both excessive inflammation and glycocalyx injury in the lung endothelium.

Pulmonary hypertension with a low cardiac index requires a higher PaO2 level to avoid tissue hypoxia.

BACKGROUND AND OBJECTIVE: The optimal oxygen supplementation needed to avoid tissue hypoxia in patients with pulmonary hypertension (PH) remains unclear. This study aimed to identify the arterial oxygen tension (PaO2 ) level needed to avoid tissue hypoxia which results in a poor prognosis in patients with PH. METHODS: We retrospectively analysed the data for 1571 right heart catheterizations in patients suspected of having PH between 1983 and 2017 at our institution. Examinations were classified according to mean pulmonary arterial pressure (mPAP), cardiac index (CI) and the presence of lung disease, pulmonary arterial hypertension (PAH) or chronic thromboembolic PH (CTEPH). The PaO2 levels needed to avoid tissue hypoxia were compared in each subgroup. RESULTS: The estimated PaO2 equivalent to a mixed venous oxygen tension (PvO2 ) of 35 mm Hg (tissue hypoxia) was 63.2 mm Hg in all patients, 77.0 mm Hg in those with decreased CI (<2.5 L/min/m2 ) and 57.0 mm Hg in those with preserved CI. Multivariate regression analysis identified mPAP, CI and PaO2 to be independent predictors of extremely low PvO2 . Similar results were observed regardless of the severity of PH or the presence of lung disease, PAH or CTEPH. The PaO2 level needed to avoid tissue hypoxia was higher in patients with mild PH and decreased CI than in those with severe PH and preserved CI (70.2 vs 61.5 mm Hg). CONCLUSION: These findings indicate that a decreased CI rather than increased mPAP induces tissue hypoxia in PH. Patients with PH and decreased CI may need adjustment of oxygen therapy at higher PaO2 levels compared with patients with preserved CI.

Isolation and characterization of endothelial-to-mesenchymal transition cells in pulmonary arterial hypertension.

Endothelial-to-mesenchymal transition (EndMT) is a process in which endothelial cells lose polarity and cell-to cell contacts, and undergo a dramatic remodeling of the cytoskeleton. It has been implicated in initiation and progression of pulmonary arterial hypertension (PAH). However, the characteristics of cells which have undergone EndMT cells in vivo have not been reported and so remain unclear. To study this, sugen5416 and hypoxia (SuHx)-induced PAH was established in Cdh5-Cre/Gt(ROSA)26Sortm4(ACTB-tdTomato,EGFP)Luo/J double transgenic mice, in which GFP was stably expressed in pan-endothelial cells. After 3 wk of SuHx, flow cytometry and immunohistochemistry demonstrated CD144-negative and GFP-positive cells (complete EndMT cells) possessed higher proliferative and migratory activity compared with other mesenchymal cells. While CD144-positive and α-smooth muscle actin (α-SMA)-positive cells (partial EndMT cells) continued to express endothelial progenitor cell markers, complete EndMT cells were Sca-1-rich mesenchymal cells with high proliferative and migratory ability. When transferred in fibronectin-coated chamber slides containing smooth muscle media, α-SMA robustly expressed in these cells compared with cEndMT cells that were grown in maintenance media. Demonstrating additional paracrine effects, conditioned medium from isolated complete EndMT cells induced enhanced mesenchymal proliferation and migration and increased angiogenesis compared with conditioned medium from resident mesenchymal cells. Overall, these findings show that EndMT cells could contribute to the pathogenesis of PAH both directly, by transformation into smooth muscle-like cells with higher proliferative and migratory potency, and indirectly, through paracrine effects on vascular intimal and medial proliferation.

Long-Term Outcome of Chronic Thromboembolic Pulmonary Hypertension at a Single Japanese Pulmonary Endarterectomy Center.

BACKGROUND: Several new treatments for chronic thromboembolic pulmonary hypertension (CTEPH) have appeared in recent years, which have led to changes in the treatment algorithm. Changes in survival rates and prognostic factors, however, have not been estimated so far.Methods and Results:Two hundred and eighty patients were diagnosed with CTEPH at Chiba University Hospital between June 1986 and June 2016. Survival rate was investigated by date of treatment initiation (group 1, 1986-1998; group 2, 1999-2008; group 3, 2009-2016). Survival rates were also evaluated by treatment strategy: balloon pulmonary angioplasty (BPA), pulmonary endarterectomy (PEA), and medical treatment. Group 3 had significantly better disease-specific survival than groups 1 and 2 (5-year survival: 91.9% vs. 67.1%, 77.0%, respectively). For the non-PEA (BPA+medication) strategy, group 3 had better disease-specific survival than groups 1 and 2 (5-year survival: 94.9% vs. 54.6%, 74.2%, respectively). The PEA strategy had significantly better survival than the medication strategy in groups 1 and 2, whereas no difference was observed between the BPA, PEA, and medication strategies in group 3. CONCLUSIONS: Survival in CTEPH in the recent era has significantly improved, especially in non-PEA patients. BPA and selective pulmonary vasodilators could improve survival in the non-PEA group. In the present study, no difference in survival was found between PEA and non-PEA.

Endothelial cell-related autophagic pathways in Sugen/hypoxia-exposed pulmonary arterial hypertensive rats.

Pulmonary arterial hypertension (PAH) is characterized by progressive obstructive remodeling of pulmonary arteries. However, no reports have described the causative role of the autophagic pathway in pulmonary vascular endothelial cell (EC) alterations associated with PAH. This study investigated the time-dependent role of the autophagic pathway in pulmonary vascular ECs and pulmonary vascular EC kinesis in a severe PAH rat model (Sugen/hypoxia rat) and evaluated whether timely induction of the autophagic pathway by rapamycin improves PAH. Hemodynamic and histological examinations as well as flow cytometry of pulmonary vascular EC-related autophagic pathways and pulmonary vascular EC kinetics in lung cell suspensions were performed. The time-dependent and therapeutic effects of rapamycin on the autophagic pathway were also assessed. Sugen/hypoxia rats treated with the vascular endothelial growth factor receptor blocker SU5416 showed increased right ventricular systolic pressure (RVSP) and numbers of obstructive vessels due to increased pulmonary vascular remodeling. The expression of the autophagic marker LC3 in ECs also changed in a time-dependent manner, in parallel with proliferation and apoptotic markers as assessed by flow cytometry. These results suggest the presence of cross talk between pulmonary vascular remodeling and the autophagic pathway, especially in small vascular lesions. Moreover, treatment of Sugen/hypoxia rats with rapamycin after SU5416 injection activated the autophagic pathway and improved the balance between cell proliferation and apoptosis in pulmonary vascular ECs to reduce RVSP and pulmonary vascular remodeling. These results suggested that the autophagic pathway can suppress PAH progression and that rapamycin-dependent activation of the autophagic pathway could ameliorate PAH.

Vildagliptin ameliorates pulmonary fibrosis in lipopolysaccharide-induced lung injury by inhibiting endothelial-to-mesenchymal transition.

BACKGROUND: Pulmonary fibrosis is a late manifestation of acute respiratory distress syndrome (ARDS). Sepsis is a major cause of ARDS, and its pathogenesis includes endotoxin-induced vascular injury. Recently, endothelial-to-mesenchymal transition (EndMT) was shown to play an important role in pulmonary fibrosis. On the other hand, dipeptidyl peptidase (DPP)-4 was reported to improve vascular dysfunction in an experimental sepsis model, although whether DPP-4 affects EndMT and fibrosis initiation during lipopolysaccharide (LPS)-induced lung injury is unclear. The aim of this study was to investigate the anti-EndMT effects of the DPP-4 inhibitor vildagliptin in pulmonary fibrosis after systemic endotoxemic injury. METHODS: A septic lung injury model was established by intraperitoneal injection of lipopolysaccharide (LPS) in eight-week-old male mice (5 mg/kg for five consecutive days). The mice were then treated with vehicle or vildagliptin (intraperitoneally, 10 mg/kg, once daily for 14 consecutive days from 1 day before the first administration of LPS.). Flow cytometry, immunohistochemical staining, and quantitative polymerase chain reaction (qPCR) analysis was used to assess cell dynamics and EndMT function in lung samples from the mice. RESULTS: Lung tissue samples from treated mice revealed obvious inflammatory reactions and typical interstitial fibrosis 2 days and 28 days after LPS challenge. Quantitative flow cytometric analysis showed that the number of pulmonary vascular endothelial cells (PVECs) expressing alpha-smooth muscle actin (α-SMA) or S100 calcium-binding protein A4 (S100A4) increased 28 days after LPS challenge. Similar increases in expression were also confirmed by qPCR of mRNA from isolated PVECs. EndMT cells had higher proliferative activity and migration activity than mesenchymal cells. All of these changes were alleviated by intraperitoneal injection of vildagliptin. Interestingly, vildagliptin and linagliptin significantly attenuated EndMT in the absence of immune cells or GLP-1. CONCLUSIONS: Inhibiting DPP-4 signaling by vildagliptin could ameliorate pulmonary fibrosis by downregulating EndMT in systemic LPS-induced lung injury.

Prognostic and pathophysiological marker for patients with chronic thromboembolic pulmonary hypertension: Usefulness of diffusing capacity for carbon monoxide at diagnosis.

BACKGROUND AND OBJECTIVE: Chronic thromboembolic pulmonary hypertension (CTEPH) is a progressive disease in some patients, despite improved treatments. Microvasculopathy has been implicated in the poor outcomes of patients with CTEPH. A reduction in the diffusing capacity for carbon monoxide (DLCO ) was previously suggested to indicate microvasculopathy in CTEPH patients; therefore, we assessed DLCO /alveolar ventilation (DLCO /VA ) as a prognostic and pathophysiological marker in CTEPH. METHODS: We performed a retrospective cohort study of 214 CTEPH patients consecutively diagnosed between 1986 and 2011. After exclusion of 24 patients because of missing DLCO data or severe obstructive impairment, the mortality rates of medically treated patients classified with normal or decreased DLCO values were compared, and prognostic factors were determined. The relationship between long-term surgical outcomes and DLCO /VA was also investigated. RESULTS: Ninety-one inoperable patients were treated medically, two of whom underwent balloon pulmonary angioplasty. Ninety-nine underwent pulmonary endarterectomy. The 5-year survival rate of medically treated patients was significantly lower in patients with decreased DLCO /VA than in those with normal values (61.4% vs 90.4%, P = 0.017). Decreased preoperative DLCO /VA was associated with a smaller percent decrease in post-operative pulmonary vascular resistance, but not with the extent of proximal thrombi; these results may support our hypothesis that DLCO reflects microvascular involvement. CONCLUSION: Decreased DLCO /VA was associated with poor outcomes of medically treated CTEPH patients; and may be useful for identifying high-risk patients, potentially leading to earlier and more appropriate interventions.

Prominin-1/CD133 expression as potential tissue-resident vascular endothelial progenitor cells in the pulmonary circulation.

Pulmonary vascular endothelial cells could contribute to maintain homeostasis in adult lung vasculature. "Tissue-resident" endothelial progenitor cells (EPCs) play pivotal roles in postnatal vasculogenesis, vascular repair, and tissue regeneration; however, their local pulmonary counterparts remain to be defined. To determine whether prominin-1/CD133 expression can be a marker of tissue-resident vascular EPCs in the pulmonary circulation, we examined the origin and characteristics of prominin-1/CD133-positive (Prom1(+)) PVECs considering cell cycle status, viability, histological distribution, and association with pulmonary vascular remodeling. Prom1(+) PVECs exhibited high steady-state transit through the cell cycle compared with Prom1(-) PVECs and exhibited homeostatic cell division as assessed using the label dilution method and mice expressing green fluorescent protein. In addition, Prom1(+) PVECs showed more marked expression of putative EPC markers and drug resistance genes as well as highly increased activation of aldehyde dehydrogenase compared with Prom1(-) PVECs. Bone marrow reconstitution demonstrated that tissue-resident cells were the source of >98% of Prom1(+) PVECs. Immunofluorescence analyses revealed that Prom1(+) PVECs preferentially resided in the arterial vasculature, including the resistant vessels of the lung. The number of Prom1(+) PVECs was higher in developing postnatal lungs. Sorted Prom1(+) PVECs gave rise to colonies and formed fine vascular networks compared with Prom1(-) PVECs. Moreover, Prom1(+) PVECs increased in the monocrotaline and the Su-5416 + hypoxia experimental models of pulmonary vascular remodeling. Our findings indicated that Prom1(+) PVECs exhibited the phenotype of tissue-resident EPCs. The unique biological characteristics of Prom1(+) PVECs predominantly contribute to neovasculogenesis and maintenance of homeostasis in pulmonary vascular tissues.

Endothelial-to-mesenchymal transition in lipopolysaccharide-induced acute lung injury drives a progenitor cell-like phenotype.

Pulmonary vascular endothelial function may be impaired by oxidative stress in endotoxemia-derived acute lung injury. Growing evidence suggests that endothelial-to-mesenchymal transition (EndMT) could play a pivotal role in various respiratory diseases; however, it remains unclear whether EndMT participates in the injury/repair process of septic acute lung injury. Here, we analyzed lipopolysaccharide (LPS)-treated mice whose total number of pulmonary vascular endothelial cells (PVECs) transiently decreased after production of reactive oxygen species (ROS), while the population of EndMT-PVECs significantly increased. NAD(P)H oxidase inhibition suppressed EndMT of PVECs. Most EndMT-PVECs derived from tissue-resident cells, not from bone marrow, as assessed by mice with chimeric bone marrow. Bromodeoxyuridine-incorporation assays revealed higher proliferation of capillary EndMT-PVECs. In addition, EndMT-PVECs strongly expressed c-kit and CD133. LPS loading to human lung microvascular endothelial cells (HMVEC-Ls) induced reversible EndMT, as evidenced by phenotypic recovery observed after removal of LPS. LPS-induced EndMT-HMVEC-Ls had increased vasculogenic ability, aldehyde dehydrogenase activity, and expression of drug resistance genes, which are also fundamental properties of progenitor cells. Taken together, our results demonstrate that LPS induces EndMT of tissue-resident PVECs during the early phase of acute lung injury, partly mediated by ROS, contributing to increased proliferation of PVECs.

Coagulation-Fibrinolysis System and Postoperative Outcomes of Patients With Chronic Thromboembolic Pulmonary Hypertension.

BACKGROUND: The postoperative changes in the coagulation-fibrinolysis system and the association between the system and postoperative course of patients with chronic thromboembolic pulmonary hypertension (CTEPH) who have undergone pulmonary endarterectomy (PEA) remain unclear. METHODS AND RESULTS: Between 1986 and 2013, 117 patients (55.1±11.2 years, preoperative mean pulmonary arterial pressure 46.5±10.5 mmHg) underwent PEA, and 15 patients died during the perioperative period. We studied the association between the preoperative coagulation-fibrinolysis markers and surgical outcomes of all patients, and the long-term outcomes of the 102 survivors from the date of PEA. We also investigated the postoperative changes in coagulation-fibrinolysis markers and their association with residual pulmonary hypertension (PH) in 20 consecutive patients. Only an elevated factor VIII level was associated with perioperative death. Thrombomodulin and plasminogen values were significantly increased after PEA. Univariate logistic regression analysis revealed that D-dimer positivity at follow-up was a risk factor for residual PH. Patients with both an elevated fibrinogen level (≥291 mg/dl [median]) and decreased plasminogen activity (<100% [median]) had significantly worse disease-specific survival than the other patients (5-year disease-specific survival: 84.0% vs. 100%, respectively; P=0.0041 [log-rank test]). CONCLUSIONS: Preoperatively high fibrinogen and low plasminogen values in patients with CTEPH are associated with poor long-term postoperative outcome. PEA benefited not only the pulmonary hemodynamics but also the coagulation-fibrinolysis system of patients.

Synergistic Effects of ACE Insertion/Deletion and GNB3 C825T Polymorphisms on the Efficacy of PDE-5 Inhibitor in Patients with Pulmonary Hypertension.

BACKGROUND: The insertion/deletion (I/D) polymorphism in the angiotensin-converting enzyme gene (ACE) and the C825T polymorphism in the G-protein ß3 subunit gene (GNB3) are associated with the efficacy of phosphodiesterase-5 inhibitor (PDE-5I) in erectile dysfunction. In addition, GNB3 genotypes could be associated with clinical worsening in pulmonary hypertension (PH) treated with PDE-5I. However, no studies have described the synergistic effects of gene polymorphisms on drug efficacy in patients with PH. OBJECTIVES: We aimed to examine the effects of combined ACE/GNB3 polymorphisms on the efficacy of PDE-5I in patients with PH. METHODS: This was a retrospective uncontrolled study. Ninety patients with pulmonary arterial hypertension (PAH) or chronic thromboembolic PH (CTEPH) were treated with PDE-5I. Freedom from clinical worsening and pre- and post-treatment parameters, including the 6-min walk distance (6MWD) and serum brain natriuretic peptide (BNP) levels, were compared between patients with ACE/GNB3 II/TT and non-II/TT genotypes. RESULTS: Time to clinical worsening was significantly longer in patients with the II/TT genotype than in those with the non-II/TT genotype (5-year freedom from clinical worsening: 100 vs. 48.8%, respectively; p = 0.018), even in patients with CTEPH alone. Post-treatment 6MWD and BNP levels in patients with the II/TT genotype tended to be better than those in patients with the non-II/TT genotype. The ACE/GNB3 genotype was a significant predictor of clinical worsening, even after adjusting for pulmonary vascular resistance and 6MWD. CONCLUSIONS: ACE and GNB3 polymorphisms may synergistically influence the efficacy of PDE-5I in patients with PH.

Vascular Repair by Tissue-Resident Endothelial Progenitor Cells in Endotoxin-Induced Lung Injury.

Vascular disruption is one of the pathological hallmarks in acute respiratory distress syndrome. Bone marrow (BM)-derived circulating endothelial progenitor cells (EPCs) and lung tissue-resident EPCs have been considered to play a pivotal role in pulmonary vascular repair; however, which population is predominant in local pulmonary vasculogenesis remains to be clarified. We therefore examined the origin of EPCs participating in the regenerative process of pulmonary vascular endothelial cells (PVECs) in experimental acute respiratory distress syndrome. Lung samples from mice administered LPS intratracheally were investigated for cell dynamics and EPC functions. Quantitative flow cytometric analysis demonstrated that the number of PVECs decreased by roughly 20% on Day 1 and then recovered on Day 7 of LPS challenge. Bromodeoxyuridine-incorporation assays and immunofluorescence microscopy demonstrated that proliferating PVECs preferentially located in the capillary vessels. Experiments using BM chimera mice revealed that most of the regenerating PVECs were tissue-resident cells, and BM-derived cells hardly engrafted as PVECs. The population of circulating putative phenotypical EPCs decreased during the first week after LPS challenge. The regenerating PVECs were characterized by high colony-forming and vasculogenic capacities, intracellular reactive oxygen species scavenging and aldehyde dehydrogenase activites, and enhanced gene expression of Abcb1b (a drug-resistant gene), suggesting that the population of PVECs included tissue-resident EPCs activated during regenerative process of PVECs. The proliferating PVECs expressed CD34, Flk-1/KDR, and c-kit more strongly and Prom1/CD133 less strongly on the surface than nonproliferating PVECs. Our findings indicated that lung tissue-resident EPCs predominantly contribute to pulmonary vascular repair after endotoxin-induced injury.

Hypoxia-induced proliferation of tissue-resident endothelial progenitor cells in the lung.

Exposure to hypoxia induces changes in the structure and functional phenotypes of the cells composing the pulmonary vascular wall from larger to most peripheral vessels. Endothelial progenitor cells (EPCs) may be involved in vascular endothelial repair. Resident EPCs with a high proliferative potential are found in the pulmonary microcirculation. However, their potential location, identification, and functional role have not been clearly established. We investigated whether resident EPCs or bone marrow (BM)-derived EPCs play a major role in hypoxic response of pulmonary vascular endothelial cells (PVECs). Mice were exposed to hypoxia. The number of PVECs transiently decreased followed by an increase in hypoxic animals. Under hypoxic conditions for 1 wk, prominent bromodeoxyuridine incorporation was detected in PVECs. Some Ki67-positive cells were detected among PVECs after 1 wk under hypoxic conditions, especially in the capillaries. To clarify the origin of proliferating endothelial cells, we used BM chimeric mice expressing green fluorescent protein (GFP). The percentage of GFP-positive PVECs was low and constant during hypoxia in BM-transplanted mice, suggesting little engraftment of BM-derived cells in lungs under hypoxia. Proliferating PVECs in hypoxic animals showed increased expression of CD34, suggesting hypoxia-induced gene expression and cell surface antigen of EPC or stem/progenitor cells markers. Isolated PVECs from hypoxic mice showed colony- and tube-forming capacity. The present study indicated that hypoxia could induce proliferation of PVECs, and the origin of these cells might be tissue-resident EPCs.

Effects of Surgical and Medical Treatment on Quality of Life for Patients With Chronic Thromboembolic Pulmonary Hypertension.

BACKGROUND: This study aimed to investigate the predictors of quality of life (QOL) in patients with chronic thromboembolic pulmonary hypertension (CTEPH), changes in QOL after surgical and medical treatments, and the relationship between baseline QOL and survival. METHODS AND RESULTS: QOL was measured in 128 patients with CTEPH (male/female: 42/86, age: 56±12 years, surgical/medical: 65/63) using the Short-Form 36 (SF-36) questionnaire. Multiple regression analysis showed pulmonary vascular resistance (PVR) and 6-min walking distance (6MWD) were associated with physical functioning (PF) (P<0.01) and physical component summary (PCS) (P<0.01). In the surgical group, 7 subscales and 2 summary scores improved significantly, and in the medical group 6 subscales and the mental component summary, although the change in QOL was greater in the surgical group. The patients in the conventional therapy group with higher PF had significantly better survival than those with lower PF (5-years survival: 89.5% vs. 50.8%, P=0.002). This difference in survival was not observed in the group receiving pulmonary arterial hypertension (PAH)-specific therapy (100% vs. 100%, P=0.746). CONCLUSIONS: PVR and 6MWD were associated with PF or PCS in CTEPH patients. QOL improved after surgical or medical therapy, with a greater change in the surgical group. PAH-specific therapy improved survival in patients with lower PF at diagnosis.

Association of plasma fibrinogen and plasminogen with prognosis of inoperable chronic thromboembolic pulmonary hypertension.

BACKGROUND: It is unclear whether abnormalities of coagulation or fibrinolysis are associated with disease progression of chronic thromboembolic pulmonary hypertension (CTEPH). The aim of this study was to investigate the association of these factors with the severity and prognosis of CTEPH. METHODS AND RESULTS: Between 1986 and 2011, plasma fibrinogen and plasminogen were measured in 89 of 106 consecutive patients with inoperable CTEPH (17 men; mean age, 55.9±14.1 years old; mean pulmonary arterial pressure, 44.0±12.4 mmHg) and the association of level with severity and prognosis were also examined. Seventeen patients had high fibrinogen and low plasminogen (medians, ≥291 mg/dl and <101%, respectively). These patients had significantly lower cardiac index (2.26±0.68 vs. 2.70±0.57 L·min(-1)·m(-2), P=0.007), higher pulmonary vascular resistance (PVR; 13.29±7.54 vs. 9.15±4.14 Wood units, P=0.003), and poor survival (5-year survival, 35.3% vs. 88.0%, P<0.001) compared to the other 72 patients. Additional analysis showed significantly poor survival in these patients compared with the other patients who did not have modern therapy. On multivariate analysis plasma fibrinogen, plasminogen and PVR were independent predictors of survival in medically treated patients. CONCLUSIONS: High plasma fibrinogen and low plasminogen are associated with poor survival in CTEPH patients without modern therapy.

Improved survival in medically treated chronic thromboembolic pulmonary hypertension.

BACKGROUND: The surgical indication for chronic thromboembolic pulmonary hypertension (CTEPH) has been modified due to recognition of peripheral type CTEPH and changes in surgical methods and skill. Bosentan and sildenafil are used as modern oral therapy (mod Tx) in patients with inoperable CTEPH, although it remains unknown whether they have positive effects on survival. METHODS AND RESULTS: A total of 202 patients were diagnosed with CTEPH at Chiba University Hospital between 1986 and 2010, 100 of whom underwent pulmonary endarterectomy. Seven medically treated patients with pulmonary vascular resistance (PVR) ≤ 300 dyn·s·cm(-5) were regarded as having mild disease. Survival rate was stratified by date of diagnosis (group 1, 1986-1998; group 2, 1999-2004; group 3, 2005-2010), and prognostic factors in the remaining 95 medically treated patients were investigated. Group 3 included the most patients treated with mod Tx (group 1, 9.1%; group 2, 24.2%; group 3, 65.0%) and had significantly better survival than either group 1 or 2 (5-year survival: group 1, 54.6%; group 2, 69.7%; group 3, 87.3%). Patients receiving mod Tx had significantly better survival than those not on mod Tx (5-year survival: 88.9% vs. 60.2%). Multivariate analysis showed that mod Tx, lower PVR, and lack of comorbidity were significant predictors of better outcome. CONCLUSIONS: Medically treated patients with CTEPH had a better survival rate, and the use of mod Tx contributed to improved survival.

Cell Tracking Suggests Pathophysiological and Therapeutic Role of Bone Marrow Cells in Sugen5416/Hypoxia Rat Model of Pulmonary Arterial Hypertension.

BACKGROUND: The mechanism of vascular remodelling in pulmonary arterial hypertension (PAH) remains unclear. Hence, defining the origin of cells constituting intractable vascular lesions in PAH is expected to facilitate therapeutic progress. Herein, we aimed to evaluate the origin of intractable vascular lesions in PAH rodent models via bone marrow (BM) and orthotopic lung transplantation (LT). METHODS: To trace BM-derived cells, we prepared chimeric rats transplanted with BM cells from green fluorescent protein (GFP) transgenic rats. Male rats were transplanted with lungs obtained from female rats and vice versa. Pulmonary hypertension was induced in the transplanted rats via Sugen5416 treatment and subsequent chronic hypoxia (Su/Hx). RESULTS: In the chimeric Su/Hx models, GFP-positive cells were observed in the pulmonary vascular area. Moreover, the right ventricular systolic pressure was significantly lower compared with wild-type Su/Hx rats without BM transplantation (P = 0.009). PAH suppression was also observed in rats that received allograft transplanted BM transplantation. In male rats that received LT and Su/Hx, BM-derived cells carrying the Y chromosome were also detected in neointimal occlusive lesions of the transplanted lungs received from female rats. CONCLUSIONS: BM-derived cells participate in pulmonary vascular remodelling in the Su/Hx rat model, whereas BM transplantation may contribute to suppression of development of PAH.

Nocturnal Hypoxemia and High Circulating TNF-α Levels in Chronic Thromboembolic Pulmonary Hypertension.

Objective Chronic thromboembolic pulmonary hypertension (CTEPH) is a form of pulmonary hypertension caused by persistent thromboemboli of the pulmonary arteries, and one of its etiological factors may be inflammation. Sleep disordered breathing (SDB) is reportedly an important complication of pulmonary hypertension. However, the association between SDB and inflammation in CTEPH has been undefined. This prospective observational study analyzed the association between the severity of SDB, pulmonary hemodynamic parameters and the systemic inflammation level in patients with CTEPH. Methods CTEPH patients admitted for a right heart catheter (RHC) examination were consecutively enrolled from November 2017 to June 2019 at the pulmonary hypertension center in Chiba University Hospital. Patients with idiopathic pulmonary arterial hypertension (IPAH) were also enrolled as a control group. All patients underwent a sleep study using a WatchPAT 200 during admission. Results The CTEPH patients showed worse nocturnal hypoxemia, oxygen desaturation index (ODI), and apnea-hypopnea index than the IPAH patients. Among these factors, only the nocturnal mean percutaneous oxygen saturation (SpO2) was negatively correlated with the pulmonary hemodynamic parameters. The circulating tumor necrosis factor-alpha (TNF-α) level was also high in the CTEPH group, and a multivariate analysis showed that the nocturnal mean SpO2 was the most important predictive factor for a high TNF-α level. Conclusion We showed that CTEPH patients had high serum TNF-α levels and that the nocturnal mean SpO2 was a predictive factor for serum TNF-α levels. Further investigations focused on nocturnal hypoxemia and the TNF-α level may provide novel insight into the etiology and new therapeutic strategies for CTEPH.

Characteristics of Japanese elderly patients with pulmonary arterial hypertension.

Previous nationwide Japanese data suggested that pulmonary arterial hypertension (PAH) predominantly affects young women. However, the number of elderly patients diagnosed with PAH has been increasing in western countries. There have been no reports on elderly PAH patients in Asian countries. This study aimed to investigate the clinical characteristics of elderly PAH patients in a Japanese cohort. Idiopathic/heritable PAH (I/H-PAH) was included in the national research project on intractable diseases. The patients were required to submit a clinical research form completed by their attending physicians. We analyzed the characteristics of Japanese I/H-PAH using the newly registered forms in 2013 (Study 1, n = 148). Also, we did a retrospective, observational cohort study at Chiba University Hospital (Study 2, n = 42). We compared the characteristics of elderly PAH patients (≥65 years old) with younger patients (<65) in both studies. Study 1 revealed a predominance of males (51% male), better hemodynamics and poorer exercise capacity in the elderly group (n = 72), compared with the younger group (n = 76) in study 1. In Study 2, elderly patients showed a male predominance (63% male), a higher ratio of smokers, a lower % carbon monoxide diffusing capacity, and poorer exercise tolerance. Elderly patients in Study 2 showed less improvement in hemodynamics with therapy. There was no significant difference in disease-specific survival between elderly and younger patients. Japanese elderly patients with I/H-PAH showed poorer exercise capacity and impaired gas exchange, but better pulmonary hemodynamics than younger patients.

Protective role of endothelial progenitor cells stimulated by riociguat in chronic thromboembolic pulmonary hypertension.

BACKGROUND: Pulmonary endothelial damage has a negative impact on the maintenance of normal pulmonary vascular function. Such damage results in delayed thrombus dissolution and vascular remodeling in chronic thromboembolic pulmonary hypertension (CTEPH). Although endothelial progenitor cells (EPCs) may be incorporated into neovasculature during vascular repair, their function in CTEPH remains unclarified, especially under the augmentation of soluble guanylate cyclase (sGC) activity. METHODS AND RESULTS: We evaluated the effect of EPCs on endothelial function and compared the effect of riociguat, a sGC stimulator, on the number and function of circulating EPCs in two groups of CTEPH patients. The two groups consisted 16 CTEPH patients who were treatment naïve (Naïve group), and 14 CTEPH patients who were being treated with riociguat, a sGC stimulator (Riociguat group). The number of circulating EPCs in the Riociguat group was significantly higher than that in the Naïve group. Gene expression levels associated with angiogenesis were significantly higher in EPCs of the Riociguat group. EPC-stimulated tube formation and migration of human pulmonary microvascular endothelial cell (hPMVEC) in the Riociguat group exceeded that in the Naïve group. The angiogenic ability of hPMVECs stimulated by EPCs in the Riociguat group was enhanced compared to that of the sGC stimulator, BAY 41-2272. CONCLUSION: These findings indicate that riociguat may induce EPCs to play a protective role via modulation of endothelial functions associated with CTEPH. TRANSLATION ASPECT OF THE WORK: Endothelial dysfunction exacerbates CTEPH. Riociguat enhanced the protective role of EPCs via neovascularization, which prevented vascular remodeling and alleviated CTEPH.