Role of Purine-Converting Ecto-Enzymes in Angiogenic Phenotype of Pulmonary Artery Adventitial Vasa Vasorum Endothelial Cells of Chronically Hypoxic Calves - Extracellular ATP and Adenosine as Regulators of Endothelial Cell Function - page 73

resembles the vascularization of tumors, as well as, the neovascularization of the
vessel wall during atherosclerotic plague development [58, 60]. These observations
raise questions as to how hypoxic conditions contribute to this neovascularization
and the process of structural remodeling of the pulmonary circulation. While many
factors with angiogenic potential have been identified, endogenous molecular fac-
tors specifically involved in hypoxia-induced expansion of the vasa vasorum, as well
as the precise cellular and molecular mechanisms contributing to this process, are
not completely understood.
5.4 Cultured PA Adventitial Vasa Vasorum Endothelial Cells
is a Novel In Vitro Angiogenic Model System
The in vitro studies of angiogenic responses in microvascular endothelial cells are
limited due to the difficulties of isolation and maintenance these cells in in vitro
conditions. We have successfully established a culture of vasa vasorum endothelial
cells (VVEC) from the adventitia of PA of neonatal calves, thereby presenting a
physiologically relevant cell system to evaluate angiogenic effects of extracellular
ATP and another nucleotides. To our knowledge, the use of this specific microvascu-
lar endothelial cell population has not been previously documented. VVEC cultures
demonstrate two distinct advantages of using the hypoxic calf model: first, after
two weeks under hypoxic conditions these animals develop pulmonary hyperten-
sion accompanied by remarkable pulmonary vascular remodeling, (similar to those
observed in human) and second, large amounts of adventitial tissues are avail-
able, which allows large scale cell isolation. Using explant techniques followed by
cloning ring differential trypsinization, VVECs can be successfully isolated from PA
adventitial compartment. These cells express CD31, Flk-1, Tie-1, vWf and eNOS,
as do pulmonary artery (Fig. 5.1) endothelial cells, cultured from the same vessel.
Fig. 5.2 Vasa vasorum endothelial cells (VVEC) can be isolated from the pulmonary artery adven-
titia and perpetuated in culture. ( a ) PA adventitial regions that develop massive neovascularization
are used for VVEC isolation; ( b ) Co-culture of VVEC and adventitial fibroblasts. Pure endothelial
cell culture can be obtained as a result of trypsinization by using cloning rings. VVEC exhibit a
“cobblestone” morphology, a characteristics of endothelial phenotype