Assessment of the role of pericyte loss in the pathophysiology of pulmonary arterial hypertension by organ-on-chip technology

Principle Investigator

Scientific interest within the context of the graduate college:

Pulmonary arterial hypertension is a cardiopulmonary disease that is characterized by profound remodeling of the pulmonary vascular tree and has a poor prognosis if left untreated. Current therapeutic approaches rely on vasodilatory drugs and do not address the underlying cause of vascular remodeling. Investigating the pathophysiology of pulmonary arterial hypertension on a novel, in vitro microvasculature-on-chip model allows to track changes of the vascular system and the dynamics of individual cell types specifically in a temporal and spatial context that was not possible before. As such, microvasculature-on-chip models – in combination with advanced imaging modalities, state-of-the-art transcriptomic and proteomic analyses, and functional read-outs – open up unprecedented avenues for the discovery of new disease mechanisms and therapeutic targets.

Project description:

Introduction: Pulmonary arterial hypertension is caused by extensive pulmonary vascular remodeling that results in increased pulmonary vascular resistance and pulmonary arterial pressure, eventually causing right ventricular dysfunction and, ultimately, right ventricular failure and death. Pulmonary vascular remodeling in pulmonary arterial hypertension is characterized by proliferation and hypertrophy of endothelial and smooth muscle cells in pulmonary resistance vessels, and a parallel loss of pulmonary microvessels, especially in the alveolar capillary network, termed microvascular rarefaction or pruning.

Recently, this microvascular rarefaction has been linked to a loss of microvascular pericytes. Pericytes are perivascular cells that are encased within the microvascular basement membrane and assist in the maturation and stabilization of microvascular networks. To this end, pericytes communicate with endothelial cells by both direct physical contact, via gap junctions and by paracrine signaling. This interaction promotes the stabilization of endothelial cells and the maintenance of vascular barrier function. Accordingly, loss or detachment of pericytes may result in increased microvascular leak and the disintegration of microvascular networks. Recent histological analyses in lung tissue samples from PAH patients and animal models of pulmonary arterial hypertension reported an increased density of pericytes in pulmonary arterial resistance vessels, while the abundance of pericytes in the pulmonary microvasculature was markedly reduced. Based on this finding, we and others have hypothesized that in PAH, pericytes may migrate from the pulmonary capillary bed to the proximal arteries where they may integrate into the arterial media as contractile cells. Such a process would not only promote arterial remodeling and muscularization, but may also in parallel destabilize the pulmonary microvasculature causing microvasculature rarefaction. The actual effect of pericyte loss on pulmonary microvascular networks has, however, so far not been elucidated due to the lack of appropriate models and methods.

Histopathological analyses in human tissue samples or animal models are typically limited to a single time point and as such, cannot assess the dynamics and interactions of distinct cell types over time and space, or track the process of microvascular rarefaction over time. In vitro assays, on the other hand, often fail to reflect the complex multicellular and physicochemical context of the intact lung. In the present project, we will apply a unique microvasculature-on-chip platform that we have successfully developed in close collaboration with microphysiological model experts in Switzerland for the study of pericyte-endothelial cell interactions in pulmonary microvascular networks, and that we will employ here to study the effects of a targeted loss of pericytes on pulmonary microvascular network stability and the associated lung endothelial phenotype. Specifically, we aim to realize the following research aims:

Aim 1: To establish a system for targeted pericyte loss in pulmonary microvasculatures-on-a-chip. Pericyte loss will be induced by batrachotoxin, an activator of voltage-gated Na⁺ channels that are present in pericytes yet not in endothelial cells. As such, batrachotoxin should cause sustained membrane depolarization and ultimately, cell death only in pericytes. Induction of selective pericyte cell death will be assayed in cultured human lung pericytes and pulmonary microvascular endothelial cells, as well as in our pulmonary microvasculatures-on-a-chip platform.

Aim 2: To study the effects of targeted pericyte loss on microvascular structure in a pulmonary microvasculatures-on-a-chip platform. To this end, selective pericyte cell death will be induced by batrachotoxin in pulmonary microvasculatures-on-a-chip generated from cultured human lung pericytes and pulmonary microvascular endothelial cells, and effects on microvasculature network morphology and function will be studied by real-time imaging and digital image analysis.

Aim 3: To study the effects of targeted pericyte loss on the transcriptomic and functional phenotype of microvascular endothelial cells in a pulmonary microvasculatures-on-a-chip platform. Following targeted pericyte loss, endothelial cells will be retrieved from pulmonary microvasculatures-on-a-chip and undergo transcriptomic analysis by single-cell RNA sequencing and phenotypic characterization.

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