The vascular adventitia acts as a biological processing center for the retrieval, integration, storage, and release of key regulators of vessel wall function. It is the most complex compartment of the vessel wall and is comprised of a variety of cells including fibroblasts, immunomodulatory cells (dendritic and macrophages), progenitor cells, vasa vasorum endothelial cells and pericytes, and adrenergic nerves. In response to vascular stress or injury, resident adventitial cells are often the first to be activated and re-programmed to then influence tone and structure of the vessel wall, to initiate and perpetuate chronic vascular inflammation, and to act to stimulate expansion of the vasa vasorum, which can act as a conduit for continued inflammatory and progenitor cell delivery to the vessel wall. This review presents the current evidence demonstrating that the adventitia acts as a key regulator of vascular wall function and structure from the “outside-in.”

An increasing volume of experimental data indicates that the adventitial fibroblast, in both the pulmonary and systemic circulations, is a critical regulator of vascular wall function in health and disease. A rapidly emerging concept is that the vascular adventitia acts as biological processing center for the retrieval, integration, storage, and release of key regulators of vessel wall function. In response to stress or injury, resident adventitial cells can be activated and reprogrammed to exhibit different functional and structural behaviors. In fact, under certain conditions, the adventitial compartment may be considered the principal injury-sensing tissue of the vessel wall. In response to vascular stresses such as overdistension and hypoxia, the adventitial fibroblast is activated and undergoes phenotypic changes, which include proliferation, differentiation, upregulation of contractile and extracellular matrix proteins, and release of factors that directly affect medial smooth muscle cell tone and growth and that stimulate recruitment of inflammatory and progenitor cells to the vessel wall. Each of these changes in fibroblast phenotype modulates either directly or indirectly changes in overall vascular function and structure. The purpose of this review is to present the current evidence demonstrating that the adventitial fibroblast acts as a key regulator of pulmonary vascular function and structure from the “outside-in.”

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, and pyridoxalphosphate-6-azophenyl-2,-4-disulfonic acid (100 M) as well as apyrase (5 units/ml) attenuated hypoxia-and ATP-induced and DNA synthesis, indicating activation and a functional role of purinoceptors under hypoxic conditions. ATP-induced DNA synthesis was augmented by hypoxia in an additive fashion, whereas ATP and hypoxia synergistically increased growth factor-induced DNA synthesis, again suggesting that ATP and hypoxia utilize similar signaling pathways to induce proliferation. Indeed, we found that ATP (100 M) and hypoxia (3% O 2 ) induced expression and activation of Egr-1 transcription factor, and both stimuli acted, in part, through a G␣ i /ERK1/2-dependent signaling pathway. Suramin, Cibacron blue 3GA, and apyrase attenuated hypoxia-induced ERK1/2 activation and Egr-1 expression. We conclude that hypoxia induces ATP release from endothelial cells and fibroblasts and that the activation of P2 purinoceptors is involved in the regulation of DNA synthesis by fibroblasts under hypoxic conditions. Hypoxia has been shown to act as a direct proliferative stimulus for fibroblasts in a variety of organs. This capability of fibroblasts is unusual, at least among mesenchymally derived cells, and appears to be important in normal development, wound healing, and fibrosis as well as in the vascular changes that characterize hypoxic pulmonary hypertension (1-3). With regard to pulmonary artery adventitial fibroblasts, we have shown that among the resident vascular wall cells they exhibit the earliest and most dramatic responses to hypoxic exposure in vivo (4). In tissue culture we have also demonstrated that hypoxia in the absence of exogenous mitogens induces proliferation of pulmonary artery fibroblasts as well as some subpopulations of aortic adventitial fibroblasts. This response is due in large part to G␣ i/o -mediated activation of a complex network of mitogen-activated protein kinases, whose specific contributions to hypoxia-induced proliferation differ from those of serum-induced growth signals (5). It remains unclear, however, whether either activation or augmentation of the hypoxiainduced growth response is due at least in part to autocrine/ paracrine responses to factors secreted by fibroblasts during hypoxia, which act through G-protein-coupled pathways.One factor that could contribute to such an autocrine loop is ATP. Purines and pyrimidines (mainly ATP, ADP, adenosine, and UTP) have widespread and specific extracellular signaling actions in the regulation of a variety of functions in many tissues and appear to have key roles in development, proliferation, differentiation, and release of hormones, neurotransmitters, and cytokines (6 -10). It is also becoming evident that alterations in the physiology of purinergic signaling may result in the development of a variety of pathologies including disorders of the immune system, neurodegenerative, and vascular diseases (7). Extracellular ATP can, in fact, stimulate the growth and proliferation of vascular smooth muscle cells (SMC), 1 and this response...

Expansion of the vasa vasorum network has been observed in a variety of systemic and pulmonary vascular diseases. We recently reported that a marked expansion of the vasa vasorum network occurs in the pulmonary artery adventitia of chronically hypoxic calves. Since hypoxia has been shown to stimulate ATP release from both vascular resident as well as circulatory blood cells, these studies were undertaken to determine if extracellular ATP exerts angiogenic effects on isolated vasa vasorum endothelial cells (VVEC) and/or if it augments the effects of other angiogenic factors (VEGF and basic FGF) known to be present in the hypoxic microenvironment. We found that extracellular ATP dramatically increases DNA synthesis, migration, and rearrangement into tube-like networks on Matrigel in VVEC, but not in pulmonary artery (MPAEC) or aortic (AOEC) endothelial cells obtained from the same animals. Extracellular ATP potentiated the effects of both VEGF and bFGF to stimulate DNA synthesis in VVEC but not in MPAEC and AOEC. Analysis of purine and pyrimidine nucleotides revealed that ATP, ADP and MeSADP were the most potent in stimulating mitogenic responses in VVEC, indicating the involvement of the family of P2Y1-like purinergic receptors. Using pharmacological inhibitors, Western blot analysis, and Phosphatidylinositol-3 kinase (PI3K) in vitro kinase assays, we found that PI3K/Akt/mTOR and ERK1/2 play a critical role in mediating the extracellular ATP-induced mitogenic and migratory responses in VVEC. However, PI3K/Akt and mTOR/p70S6K do not significantly contribute to extracellular ATP-induced tube formation on Matrigel. Our studies indicate that VVEC, isolated from the sites of active angiogenesis, exhibit distinct functional responses to ATP, compared to endothelial cells derived from large pulmonary or systemic vessels. Collectively, our data support the idea that extracellular ATP participates in the expansion of the vasa vasorum that can be observed in hypoxic conditions.