“Primary Pulmonary Hypertension has been called the cardiologist’s cancer”
Greg Elliott in personal conversation with Norbert Voelkel, early 80’s:
Pulmonary Arterial Hypertension (PAH) can be a rapidly progressive and devastating disease characterized by dysfunction and remodeling of the pulmonary vasculature, leading to increased pulmonary vascular resistance. The increased vascular resistance pushes the right ventricle (RV) into adaptive compensatory remodeling by hypertrophy, but eventually RV dilatation, heart failure and death of the patient become inevitable [1, 2]. Today, 3 pathways are targeted in PAH treatment: the nitric oxide-cyclic guanosine monophosphate pathway, the endothelin pathway and the prostacyclin pathway . Although treatments affecting these pathways delay disease progression and increase survival rates [3, 4], they do not cure PAH . The pathogenic paradigm of PAH has shifted from pulmonary vasoconstriction driven by smooth muscle cells to vascular remodeling affecting all vessel wall layers. In this chapter, a brief overview will be given of how PAH research has converged on the role of the endothelial cell (EC). It will be argued that the development of new animal models has been critical to the development of a new PAH paradigm.
In 1970, the first morphologic characterization of the pulmonary hypertensive lung was published by Wagenvoort et al. [5, 6], followed by the first WHO classification of Pulmonary Hypertension in 1972. In this classification, the pathological focus in PAH, then called Primary Pulmonary Hypertension, was on the vascular media of the pulmonary arterioles, featuring hypertrophied and hyperplastic pulmonary artery smooth muscle cells (PASMC) [1, 2, 7]. The “vascular media paradigm” of PAH ascribed a major pathogenic role to sustained pulmonary vasoconstriction and was fueled by two PAH outbreaks related to the use of the appetite suppressants Aminorex (Menocil®) and fenfluramine (Ponderal®) in the late 60’s and 80’s. Both drugs are serotonin transporter substrates acting on the PASMC and promote vasoconstriction and PASMC hypertrophy and proliferation. At that time, the signature plexiform lesion was regarded as an epiphenomenon.
The description of idiopathic PAH as ‘the cardiologist’s cancer’ in the early 80’s meant at that time PAH and cancer shared a common clinical and scientific context. Both conditions were untreatable due to a profound lack of understanding and absence of technologies to study pathogenesis and pathobiology. As dedicated research did not lead to a medical cure for PAH, the idea that other vascular wall cells than smooth muscle cells were also involved in the disease was gradually accepted. For a better understanding of PAH, it was necessary to study the cellular and molecular aspects of the disease. An expanding research effort revealed the mechanisms by which PAH is characterized today: endothelial dysfunction, PASMC hypertrophy and hyperplasia, persistent inflammation and dysimmunity and dysregulated intra- and extracellular cell signaling in the cells of all layers of the pulmonary vessel bed, leading to pulmonary vessel remodeling [8–10]. Endothelial dysfunction is reflected in increased activity of contractive vasoactive agents as endothelin-1, serotonin, angiotensin II and decreased activity of dilative vasoactive agents as nitric oxide and prostacyclin, felt to lead to persistent vasoconstriction. The hyperproliferative endothelium in PAH disobeys the “law-of-the-monolayer” and has a monoclonal origin in plexiform lesions [11–13]. This hyperproliferation is mediated by increased activity of growth factors as fibroblast growth factor-2, platelet derived growth factor and epidermal growth factor and is accompanied by apoptopic resistance [14–17]. Also in the PASMC the increased activity of growth factors and apoptosis resistance are observed. As well in the EC, hyperproliferation of PASMC is mediated by migration and dysregulated BMPRII signaling, for example due to the germline BMPRII-mutation, which has a penetrance of approximately 75% in hereditary familial PAH [10, 14, 15]. Another germline mutation in PAH is found in the KNCK3 gene, which decreases the activity of TWIK-related acid-sensitive K+ channel-1 . Together with alterations in the voltage-gated K+ channel, transient receptor potential 1 and 6 and calcium sensing receptor, the PASMC is hyperpolarized [9, 18, 19]. Moreover, the PASMC in PAH goes through an energetic shift to glycolysis , also found in fibroblast of the pulmonary adventitia , which is similar to the Warburg effect. These dysregulated mechanisms also affect the extracellular matrix , and contribute to the abnormal cell signaling leading to disordered angiogenesis [16, 17, 23], and recruit inflammatory cells in the pulmonary vasculature which can release cytokines and chemokines as for example interleukin 1 and 6 [11, 15, 24–26]. It is now recognized that all layers of the vessel wall contribute to the pulmonary vascular remodeling; including the hyperproliferative endothelium, and the plexiform lesion is no longer neglected as a hallmark of PAH [27, 28]. Many of the observations made during the study of pulmonary vascular remodeling in PAH, echoed the description of the hallmarks of cancer as described by Hanahan and Weinberg [28–31]. This expansion in knowledge gave rise to the exploration of potential treatment targets, which were already studied for their efficacy in cancer, such as Histone DeACetylase inhibitors (HDACis) and Tyrosine Kinase Inhibitors (TKIs) .
Experimental models to resemble pulmonary arterial hypertension
Translational research on the development of pulmonary hypertension and potential treatments of the disease has relied on animal models that replicate one or more important aspects of the disease [33–36]. Reeves and Grover were the first to use hypoxia induced pulmonary hypertension as a model of the human condition when they studied Brisket disease in cattle kept at high altitude [37, 38]. Exposure to hypoxia was soon also used to study pulmonary hypertension in rats. In addition, an animal model based on the administration of monocrotaline was first used in 1967 [39, 40]. The monocrotaline and chronic hypoxia animal models of pulmonary hypertension both exhibit profound remodeling of the vascular media and hypertrophy of PASMC. Because these phenomena were considered singularly important in human pulmonary hypertension, it was felt that there was little need to develop alternative animal models.
The expansion in pathobiological knowledge of PAH has coincided with the establishment of new animal models of the disease. When in 2001 the Sugen Hypoxia model (SuHx) of PAH was discovered , the striking resemblance was appreciated between the pathological changes in the SuHx rat and the human PAH lung. More specifically, it was noted that the intima remodeling and obliterative vascular remodeling that is typical of human PAH, was mimicked by the lung vascular changes found in the SuHx rat [35, 41, 42]. The SuHx model also brought the understanding that the emergence of a proliferative endothelium could require an initial phase of endothelial apoptosis, as it could be prevented by a broad spectrum caspase inhibitor [41, 43]. Also in other new animal models intimal proliferation were found in the combination of monocrotaline plus aortic caval shunt or monocrotaline plus pneumonectomy [44–46].
Despite increasing popularity, the SuHx model has not (yet) become a ‘gold standard’ for animal research in PAH. In comparison to the classic animal models, the SuHx model is more complex, has not been thoroughly characterized and is more difficult to implement and harmonize among laboratories. In addition, a vast amount of translational studies has been performed with the chronic hypoxia and monocrotaline models and this body of work is a heritage of historical literature to which new data is conveniently compared. Therefore, many translational studies are still performed using the traditional animal models. Remarkably, many potential treatments adopted from the cancer field, are still explored in the classic animal models that do not represent the pathobiology of angio-obliterative PAH nor offer a hyperproliferative endothelium as a treatment target. This might explain why therapeutic interventions seem curative in animal studies whereas human PAH remains refractory to treatment [47, 48].
Due to the ineffectiveness of vasodilating drugs and aided by many cellular and molecular findings, the pathogenic paradigm of PAH has shifted from vasoconstriction towards vessel remodeling with resemblances to malignancy. Concurrently, novel animal models have evolved, which feature the human hallmark of PAH, a hyperproliferative endothelium, that is not found in more traditional animal models. Therefore, when these traditional animal models are used to assess anti-proliferative drug targets of the endothelium, they become unpredictable in their translational value. Of course, due to the unknown pathogenesis of PAH, it is paradoxically true that no animal model can resemble PAH completely. However, with the increasing realization and evaluation of the different pathobiologies resembled by animal models [33–35], the selection of the used animal model should be explicitly motivated.
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