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Most published studies addressing the role of hypoxia inducible factors (HIFs) in hypoxia-induced pulmonary hypertension development employ models that may not recapitulate the clinical setting, including the use of animals with pre-existing lung/vascular defects secondary to embryonic HIF ablation or activation. Furthermore, critical questions including how and when HIF signalling contributes to hypoxia-induced pulmonary hypertension remain unanswered.Normal adult rodents in which global HIF1 or HIF2 was inhibited by inducible gene deletion or pharmacological inhibition (antisense oligonucleotides (ASO) and small molecule inhibitors) were exposed to short-term (4 days) or chronic (4-5 weeks) hypoxia. Haemodynamic studies were performed, the animals euthanised, and lungs and hearts obtained for pathological and transcriptomic analysis. Cell-type-specific HIF signals for pulmonary hypertension initiation were determined in normal pulmonary vascular cells in vitro and in mice (using cell-type-specific HIF deletion).Global Hif1a deletion in mice did not prevent hypoxia-induced pulmonary hypertension at 5 weeks. Mice with global Hif2a deletion did not survive long-term hypoxia. Partial Hif2a deletion or Hif2-ASO (but not Hif1-ASO) reduced vessel muscularisation, increases in pulmonary arterial pressures and right ventricular hypertrophy in mice exposed to 4-5 weeks of hypoxia. A small molecule HIF2 inhibitor (PT2567) significantly attenuated early events (monocyte recruitment and vascular cell proliferation) in rats exposed to 4 days of hypoxia, as well as vessel muscularisation, tenascin C accumulation and pulmonary hypertension development in rats exposed to 5 weeks of hypoxia. In vitro, HIF2 induced a distinct set of genes in normal human pulmonary vascular endothelial cells, mediating inflammation and proliferation of endothelial cells and smooth muscle cells. Endothelial Hif2a knockout prevented hypoxia-induced pulmonary hypertension in mice.Inhibition of HIF2 (but not HIF1) can provide a therapeutic approach to prevent the development of hypoxia-induced pulmonary hypertension. Future studies are needed to investigate the role of HIFs in pulmonary hypertension progression and reversal.

Studies in various animal models suggest an important role for pulmonary macrophages in the pathogenesis of pulmonary hypertension (PH). Yet, the molecular mechanisms characterizing the functional macrophage phenotype relative to time and pulmonary localization and compartmentalization remain largely unknown. In this study, we used a hypoxic murine model of PH in combination with FACS to quantify and isolate lung macrophages from two compartments over time and characterize their programing via RNA sequencing approaches. In response to hypoxia, we found an early increase in macrophage number that was restricted to the interstitial/perivascular compartment, without recruitment of macrophages to the alveolar compartment or changes in the number of resident alveolar macrophages. Principal component analysis demonstrated significant differences in overall gene expression between alveolar and interstitial macrophages (IMs) at baseline and after 4 and 14 d hypoxic exposure. Alveolar macrophages at both day 4 and 14 and IMs at day 4 shared a conserved hypoxia program characterized by mitochondrial dysfunction, proinflammatory gene activation, and mTORC1 signaling, whereas IMs at day 14 demonstrated a unique anti-inflammatory/proreparative programming state. We conclude that the pathogenesis of vascular remodeling in hypoxic PH involves an early compartment-independent activation of lung macrophages toward a conserved hypoxia program, with the development of compartment-specific programs later in the course of the disease. Thus, harnessing time- and compartment-specific differences in lung macrophage polarization needs to be considered in the therapeutic targeting of macrophages in hypoxic PH and potentially other inflammatory lung diseases.

Macrophage accumulation is not only a characteristic hallmark but is also a critical component of pulmonary artery remodeling associated with pulmonary hypertension (PH). However, the cellular and molecular mechanisms that drive vascular macrophage activation and their functional phenotype remain poorly defined. Using multiple levels of in vivo (bovine and rat models of hypoxia-induced PH, together with human tissue samples) and in vitro (primary mouse, rat, and bovine macrophages, human monocytes, and primary human and bovine fibroblasts) approaches, we observed that adventitial fibroblasts derived from hypertensive pulmonary arteries (bovine and human) regulate macrophage activation. These fibroblasts activate macrophages through paracrine IL-6 and STAT3, HIF1, and C/EBPβ signaling to drive expression of genes previously implicated in chronic inflammation, tissue remodeling, and PH. This distinct fibroblast-activated macrophage phenotype was independent of IL-4/IL-13-STAT6 and TLR-MyD88 signaling. We found that genetic STAT3 haplodeficiency in macrophages attenuated macrophage activation, complete STAT3 deficiency increased macrophage activation through compensatory upregulation of STAT1 signaling, and deficiency in C/EBPβ or HIF1 attenuated fibroblast-driven macrophage activation. These findings challenge the current paradigm of IL-4/IL-13-STAT6-mediated alternative macrophage activation as the sole driver of vascular remodeling in PH, and uncover a cross-talk between adventitial fibroblasts and macrophages in which paracrine IL-6-activated STAT3, HIF1α, and C/EBPβ signaling are critical for macrophage activation and polarization. Thus, targeting IL-6 signaling in macrophages by completely inhibiting C/EBPβ or HIF1α or by partially inhibiting STAT3 may hold therapeutic value for treatment of PH and other inflammatory conditions characterized by increased IL-6 and absent IL-4/IL-13 signaling.

The development of parenteral prostacyclin therapy marked a dramatic breakthrough in the treatment of pulmonary arterial hypertension (PAH). Intravenous (IV) epoprostenol was the first PAH specific therapy and to date, remains the only treatment to demonstrate a mortality benefit. Because of the inherent complexities and risks of treating patients with continuous infusion IV therapy, there is great interest in the development of an oral prostacyclin analog that could mimic the benefits of IV therapy. Herein, we highlight the development of oral prostacyclin therapy, focusing on oral treprostinil, the only US Food and Drug Administration approved oral prostacyclin. Recent Phase III clinical trials have shown the drug to improve exercise tolerance in treatment-naïve PAH patients, but not patients on background oral therapy. Oral treprostinil appears to be most efficacious at higher doses, but its side effect profile and complexities with dosing complicate its use. While oral treprostinil's current therapeutic role in PAH remains unclear, ongoing studies of this class of medication should help clarify their role in the treatment of PAH. [ABSTRACT FROM AUTHOR]

Macrophage accumulation is not only a characteristic hallmark but is also a critical component of pulmonary artery remodeling associated with pulmonary hypertension (PH). However, the cellular and molecular mechanisms that drive vascular macrophage activation and their functional phenotype remain poorly defined. Using multiple levels of in vivo (bovine and rat models of hypoxia-induced PH, together with human tissue samples) and in vitro (primary mouse, rat, and bovine macrophages, human monocytes, and primary human and bovine fibroblasts) approaches, we observed that adventitial fibroblasts derived from hypertensive pulmonary arteries (bovine and human) regulate macrophage activation. These fibroblasts activate macrophages through paracrine IL-6 and STAT3, HIF1, and C/EBPβ signaling to drive expression of genes previously implicated in chronic inflammation, tissue remodeling, and PH. This distinct fibroblast-activated macrophage phenotype was independent of IL-4/IL-13-STAT6 and TLR-MyD88 signaling. We found that genetic STAT3 haplodeficiency in macrophages attenuated macrophage activation, complete STAT3 deficiency increased macrophage activation through compensatory upregulation of STAT1 signaling, and deficiency in C/EBPβ or HIF1 attenuated fibroblast-driven macrophage activation. These findings challenge the current paradigm of IL-4/IL-13-STAT6-mediated alternative macrophage activation as the sole driver of vascular remodeling in PH, and uncover a cross-talk between adventitial fibroblasts and macrophages in which paracrine IL-6-activated STAT3, HIF1a, and C/EBPβ signaling are critical for macrophage activation and polarization. Thus, targeting IL-6 signaling in macrophages by completely inhibiting C/EBPβ or HIF1a or by partially inhibiting STAT3 may hold therapeutic value for treatment of PH and other inflammatory conditions characterized by increased IL-6 and absent IL-4/IL-13 signaling. [ABSTRACT FROM AUTHOR]

Rationale: Pulmonary arterial hypertension (PAH) and chronic thromboembolic pulmonary hypertension (CTEPH) cause right ventricular dysfunction which can impact other solid organs. However, the profiles and consequences of hepatic injury due to PAH and CTEPH have not been well-studied., Objectives: We aimed to identify underlying patterns of liver injury in a cohort of PAH and CTEPH patients enrolled in 15 randomized clinical trials conducted between 1998 and 2014., Methods: We used unsupervised machine learning to identify liver injury clusters in 13 trials and validated the findings in two additional trials. We then determined whether these liver injury clusters were associated with clinical outcomes or treatment effect heterogeneity., Measurements and Main Results: Our training dataset included 4,219 patients and our validation dataset included 1,756 patients with serum total bilirubin, alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase, and albumin data. Using k-means clustering, we identified phenotypes with no liver injury, hepatocellular injury, cholestatic injury, and combined injury patterns. Patients in the cholestatic injury liver cluster had the shortest time to clinical worsening and the highest risk of mortality. The cholestatic injury group also experienced the greatest placebo-corrected treatment effect on six-minute walk distance. Randomization to the experimental arm transitioned patients to a healthier liver status., Conclusions: Liver injury was associated with adverse outcomes in patients with PAH and CTEPH. Randomization to active treatment had beneficial effects on liver health compared to placebo. The role of liver disease (often subclinical) in determining outcomes warrants prospective studies.

Hypoxic pulmonary hypertension (PH) comprises a heterogeneous group of diseases sharing the common feature of chronic hypoxia-induced pulmonary vascular remodeling. The disease is usually characterized by mild to moderate pulmonary vascular remodeling that is largely thought to be reversible compared with the progressive irreversible disease seen in World Health Organization (WHO) group I disease. However, in these patients, the presence of PH significantly worsens morbidity and mortality. In addition, a small subset of patients with hypoxic PH develop "out-of-proportion" severe pulmonary hypertension characterized by pulmonary vascular remodeling that is irreversible and similar to that in WHO group I disease. In all cases of hypoxia-related vascular remodeling and PH, inflammation, particularly persistent inflammation, is thought to play a role. This review focuses on the effects of hypoxia on pulmonary vascular cells and the signaling pathways involved in the initiation and perpetuation of vascular inflammation, especially as they relate to vascular remodeling and transition to chronic irreversible PH. We hypothesize that the combination of hypoxia and local tissue factors/cytokines ("second hit") antagonizes tissue homeostatic cellular interactions between mesenchymal cells (fibroblasts and/or smooth muscle cells) and macrophages and arrests these cells in an epigenetically locked and permanently activated proremodeling and proinflammatory phenotype. This aberrant cellular cross-talk between mesenchymal cells and macrophages promotes transition to chronic nonresolving inflammation and vascular remodeling, perpetuating PH. A better understanding of these signaling pathways may lead to the development of specific therapeutic targets, as none are currently available for WHO group III disease., (Copyright © 2015 the American Physiological Society.)