Rationale: Prostacyclin analogs, used to treat idiopathic pulmonary arterial hypertension (IPAH), are assumed to work through prostacyclin (IP) receptors linked to cyclic AMP (cAMP) generation, although the potential to signal through peroxisome proliferatoractivated receptor-g (PPARg) exists. Objectives: IP receptor and PPARg expression may be depressed in IPAH. We wished to determine if pathways remain functional and if analogs continue to inhibit smooth muscle proliferation. Methods: We used Western blotting to determine IP receptor expression in peripheral pulmonary arterial smooth muscle cells (PASMCs) from normal and IPAH lungs and immunohistochemistry to evaluate IP receptor and PPARg expression in distal arteries. Measurements and Main Results: Cell proliferation and cAMP assays assessed analog responses in human and mouse PASMCs and HEK-293 cells. Proliferative rates of IPAH cells were greater than normal human PASMCs. IP receptor protein levels were lower in PASMCs from patients with IPAH, but treprostinil reduced replication and treprostinil-induced cAMP elevation appeared normal. Responses to prostacyclin analogs were largely dependent on the IP receptor and cAMP in normal PASMCs, although in IP 2/2 receptor cells analogs inhibited growth in a cAMP-independent, PPARg-dependent manner. In IPAH cells, antiproliferative responses to analogs were insensitive to IP receptor or adenylyl cyclase antagonists but were potentiated by a PPARg agonist and inhibited (z 60%) by the PPARg antagonist GW9662. This coincided with increased PPARg expression in the medial layer of acinar arteries. Conclusions: The antiproliferative effects of prostacyclin analogs are preserved in IPAH despite IP receptor down-regulation and abnormal coupling. PPARg may represent a previously unrecognized pathway by which these agents inhibit smooth muscle proliferation.

Stable prostacyclin analogues can signal through cell surface IP receptors or by ligand binding to nuclear peroxisome proliferator-activated receptors (PPARs). So far these agents have been reported to activate PPARα and PPARδ but not PPARγ. Given PPARγ agonists and prostacyclin analogues both inhibit cell proliferation, we postulated that the IP receptor might elicit PPARγ activation. Using a dual luciferase reporter gene assay in HEK-293 cells stably expressing the IP receptor or empty vector, we found that prostacyclin analogues only activated PPARγ in the presence of the IP receptor. Moreover, the novel IP receptor antagonist, RO1138452, but not inhibitors of the cyclic AMP pathway, prevented activation. Likewise, the anti-proliferative effects of treprostinil observed in IP receptor expressing cells, were partially inhibited by the PPARγ antagonist, GW9662. We conclude that PPARγ is activated through the IP receptor via a cyclic AMP-independent mechanism and contributes to the anti-growth effects of prostacyclin analogues.