Increased expression and altered subcellular distribution of PKC-δ and PKC-ɛ in pulmonary arteries exposed to hypoxia and 15-HETE

15-Hydroxyeicosatetraenoic acid (15-HETE), a product of arachidonic acid (AA) catalyzed by 15-lipoxygenase (15-LOX), is an important mediator of hypoxic pulmonary vasoconstriction (HPV). We have previously reported that 15-HETE-induced pulmonary vasoconstriction occurs via protein kinase C (PKC) pathway, however, the role of PKC isoforms involved in 15-HETE-induced pulmonary vasoconstriction remains poorly understood. To examine the potential role of PKC-δ and PKC-ɛ isoforms that appear to be involved in 15-HETE-induced pulmonary artery (PA) contraction, a combination of immunofluorescence, western blotting, semi-quantitative PCR and functional contractile tension approaches on rat PA rings were utilized. We found that 15-HETE activates the translocation of PKC-δ and PKC-ɛ from the cytoplasm to the membranes of pulmonary arterial smooth muscle cells (PASMCs). However, the alteration was significantly reversed by nordihydroguairetic acid (NDGA), a 15-LOX inhibitor which blocked the formation of endogenous 15-HETE. Both endogenous and exogenous 15-HETE enhanced the expression of PKC-δ and PKC-ɛ in PASMCs exposure to hypoxia. The PKC inhibitor Gö6983 and rottlerin (PKC-δ selective), and the inhibitor selective for PKC-ɛ peptide significantly attenuated constriction effect of 15-HETE on isolated PA rings of rats maintained for 9 days in hypoxic environments (FiO2 = 0.12) compared with siblings rats under normoxia. Thus, these findings indicate that PKC-δ and PKC-ɛ contributing to hypoxic pulmonary artery contraction elicited by 15-HETE.

Research highlights▶ Hypoxia increase the expression of PKC-δ and PKC-ɛ, which is regulated by 15-HETE, leading to pulmonary vasoconstriction. ▶ Secondly, 15-HETE seems accountable for the PKC-δ and PKC-ɛ translocation from cytoplasm to membranes of PASMCs during hypoxia. ▶ Thirdly, 15-HETE-induced PA constriction was markedly attenuated by two separate selective PKC-δ and PKC-ɛ inhibitors. These data present strong evidence that PKC-δ and PKC-ɛ contributing to PA constriction elicited by 15-HETE.