Targeting the adventitial microenvironment in pulmonary hypertension: A potential approach to therapy that considers epigenetic change

Abstract

Experimental data indicate that the adventitial compartment of blood vessels, in both the pulmonary and systemic circulations, like the connective tissue stroma in tissues throughout the body, is a critical regulator of vessel wall function in health and disease. It is clear that adventitial cells, and in particular the adventitial fibroblast, are activated early following vascular injury, and play essential roles in regulating vascular wall structure and function through production of chemokines, cytokines, growth factors, and reactive oxygen species (ROS). The recognition of the ability of these cells to generate and maintain inflammatory responses within the vessel wall provides insight into why vascular inflammatory responses, in certain situations, fail to resolve. It is also clear that the activated adventitial fibroblast plays an important role in regulating vasa vasorum growth, which can contribute to ongoing vascular remodeling by acting as a conduit for delivery of inflammatory and progenitor cells. These functions of the fibroblast clearly support the idea that targeting chemokine, cytokine, adhesion molecule, and growth factor production in activated fibroblasts could be helpful in abrogating vascular inflammatory responses and thus in ameliorating vascular disease. Further, the recent observations that fibroblasts in vascular and fibrotic diseases may maintain their activated state through epigenetic alterations in key inflammatory and pro-fibrotic genes suggests that current therapies used to treat pulmonary hypertension may not be sufficient to induce apoptosis or to inhibit key inflammatory signaling pathways in these fibroblasts. New therapies targeted at reversing changes in the acetylation or methylation status of key transcriptional networks may be needed. At present, therapies specifically targeting abnormalities of histone deacytelase (HDAC) activity in fibroblast-like cells appear to hold promise.

Keywords: adventitial; epigenetics; fibroblasts; pulmonary hypertension.

Figure 1

Contrasting hypotheses regarding origin and perpetuation of vascular remodeling and inflammation. (A) Traditionally, remodeling and inflammation have been considered an “Inside/Out” response centered on endothelial injury, leukocyte/monocyte recruitment to the intima of blood vessels, followed by activation of medial smooth muscle cells. In this hypothesis, remodeling is largely thought to be mediated by endothelial cell activation, injury or death, abnormalities in endothelial-smooth muscle communication and resultant hyper-proliferation of either/or both cell types and recruitment of inflammatory cells, which are directed from the lumen of the pulmonary artery. (B) The “Outside-In” hypothesis suggests that vascular inflammation because of resident professional (dendritic cells, macrophages and lymphocytes) and nonprofessional (fibroblasts) immune cells occurs early and persists in the adventitia. Fibroblast activation, leukocyte and progenitor cell accumulation and retention lead to remodeling not only of the adventitia, but cause subsequent changes in the media and ultimately even the intima. Thus, the adventitia, as opposed to its′ usual depiction (Panel A) of an unimportant simple support structure, is actually a highly cellular, metabolically active, regulatory compartment of the vessel wall, capable of controlling tone, structure, and inflammation from the outside-in.

Figure 2

The activated adventitial fibroblast plays a pivotal role in the recruitment and retention of inflammatory cells in the vascular wall. In response to a variety of environmental stimuli, the adventitial fibroblast, potentially through a number of cell surface receptors, including Toll-like receptors (TLR), integrins, and receptors for advanced glycosolation end products (RAGE) is activated to produce extracellular matrix proteins, matricellular proteins, matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). In addition, the fibroblast upregulates production of chemokines and cytokines, adhesion molecules, and angiogenic factors. This leads to the recruitment of leukocytes and ultimately to an increase in vasa vasorum density or adventitial neovascularization, which perpetuates the inflammatory process.

Figure 3

Animal models of pulmonary hypertension are consistently characterized by adventitial/peri-vascular accumulation of leukocytes, and in particular mononuclear cells. (A) Illustrates the accumulation of mononuclear cells/macrophages in the adventitial/ perivascular regions of chronically hypoxic rats, calves, and mice. (B) Perivascular accumulation of mononuclear cells (CD14+ cells) is observed in the human patient with iPAH. (C) Mouse models of pulmonary hypertension, including transgenic VIP−/− mice, adiponectin (APN) deficient mice, IL-6 transgenic mice, and the BMPR2 mutated mouse model all exhibit perivascular accumulation of leukocytes.[–38]

Figure 4

Vasa vasorum expansion in the hypoxic calf model of pulmonary hypertension and in iPAH. (A) In both large (panels A and C) and small pulmonary arteries (panels B and D) of the chronically hypoxic calf, marked expansion of the vasa vasorum is observed. Panels A and B demonstrate H and E staining. Panels C and D demonstrate von Willebrand Factor expression. (B) In pulmonary arterial lesions of patients with iPAH a large expansion of CD34+ vasa vasorum vessels around the remodeled pulmonary arteries is observed (panels B, C, and D). Rare CD 34+ vessels around pulmonary arteries of control subjects are observed (panel A).[4058]

Figure 5

Fibroblasts from the chronically hypoxic pulmonary artery express an “imprinted” pro-inflammatory phenotype. Fibroblasts derived from control and hypoxic pulmonary hypertensive calves were cultured and in the absence of any exogenous stimuli, the fibroblast from pulmonary hypertensive animals (PH-Fibs) exhibited a constitutively activated “imprinted” phenotype characterized by overexpression of cytokines, chemokines and adhesion molecules. This phenotype was confirmed at the mRNA level and at the protein level both in vivo and in vitro. At the protein level IL-1β, SDF-1, MCP-1, and VECAM are dramatically upregulated both in vivo and maintain this phenotype in vitro. Confirmation of MCP-1 and VECAM are shown in the bar graphs below.[41]

Figure 6

The constitutively activated “imprinted” phenotype of fibroblasts is due, at least in part, to increased HDAC activity.

Figure 7

HDAC inhibitors attenuate the pro-liferative and -inflammatory phenotype exhibited by the constitutively activated fibroblast from pulmonary hypertensive animals.